Cell atlas of human psoriasis and epidermal specific Ube2l3 deficiency mouse highlighting CXCL16/CXCR6 orchestrating the development of psoriasis

Cell atlas of human psoriasis and epidermal specific Ube2l3 deficiency mouse highlighting CXCL16/CXCR6 orchestrating the development of psoriasis | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Article Cell atlas of human psoriasis and epidermal specific Ube2l3 deficiency mouse highlighting CXCL16/CXCR6 orchestrating the development of psoriasis Xiao-Yong Man, Xue-Yan Chen, Li-Ran Ye, Ni-Chang Fu, Si-Qi Chen, and 8 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-5973089/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 13 Oct, 2025 Read the published version in Nature Communications → Version 1 posted You are reading this latest preprint version Abstract Psoriasis is a chronic, complex immune-mediated inflammatory disorder with cutaneous and systemic manifestations in which keratinocytes, dendritic cells and T cells have central roles. UBE2L3 may be a protective biomarker that regulates the pathogenesis of psoriasis. Here, we identified the IL-17A signaling similarity between human psoriatic skin and Ube2l3 conditional knockout mouse skin in the epidermis rather than dermis. IL-17A was regulated by CXCR6 + Vγ2 + γδT in mouse while CXCR6 + CD8 + T in human. CXCL16 is the only chemokines whose bind to stimulate CXCR6. Ube2l3 reduction in keratinocytes activated IL-1β and then promote CXCL16 expression through STAT3 signaling. Up-regulated CXCL16 in keratinocytes and cDC2/mDC then attracted Vγ2 + γδT17 or CD8 + T to secrete IL-17A and form a positive feedback loop in keratinocytes supporting psoriatic lesion. Thus, UBE2L3 is a keratinocyte-intrinsic suppressor of epidermal IL-17 production in Vγ2 + γδT in mouse and CD8 + T in human through CXCL16/CXCR6 signaling pathway in psoriasis. Biological sciences/Immunology/Inflammation/Chronic inflammation Health sciences/Biomarkers/Prognostic markers Health sciences/Molecular medicine Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 INTRODUCTION Psoriasis is a chronic, complex immune-mediated inflammatory disorder with cutaneous and systemic manifestations 1 . Its pathogenesis is characterized by the involvement of keratinocytes, dendritic cells and T cells, which play a central role in the disease process 2 . To facilitate research on the mechanisms of action that drive inflammatory and autoimmune processes associated with psoriasis, it would be highly beneficial to have a suitable animal model 3 , 4 . Moreover, keratinocytes may be a crucial factor in both the onset and perpetuation of psoriasis 5 . Our and other previous research have indicated that diminished Ubiquitin Conjugating Enzyme E2 L3 (UBE2L3) expression is linked to elevated Interleukin 1β (IL-1β) production in the epidermis of psoriasis. 6 , 7 . The epidermal immune environment is of pivotal importance in the pathogenesis of psoriasis 8 . Furthermore, UBE2L3 interacts with the ubiquitin enzyme 3, including TRIP12 and AREL1, to facilitate the turnover of the precursor IL-1β (pro-IL-1β) 9 . This process is of critical importance to the functioning of the immune system. Consequently, we firstly established a mouse model of epidermal Ube2l3 deficiency in order to gain further insight into the pathogenesis of psoriasis. This mouse model offers a valuable opportunity to explore the pathogenesis of psoriasis 3 , 10 . It is of great importance to compare the mutant mice to psoriatic patient samples at the level of bulk RNA, single cell RNA, and protein. In contrast to adaptive IL-17-producing T cells, dermal IL-17-producing γδT (γδT17) cells are programmed for the earliest IL-17 response to various pathogens 11 . In these settings, γδT17 cells have been demonstrated to respond to cytokines, predominantly IL-23 and IL-1β 12 . Vγ2 T (IL-17A producers,Heilig & Tonegawa nomenclature) cells have been demonstrated to contribute to the pathogenesis of autoimmune diseases by through the production of IL-17A 13 . Integration of single cell sequencing (scRNA-seq) data and spatial-seq transcriptomic CD8 + IL17 + T (Tc17) enabled the definition of Tc17, which was identified as a principal source of IL-17A in psoriatic skin 14 , 15 . However, the function of γδT17 and Tc17 in epidermal psoriatic lesions remains to be elucidated. Chemokine (C-X-C motif) ligand 16 (CXCL16) functions as a ligand for C-X-C chemokine receptor type 6 (CXCR6), and both are all upregulated in psoriasis. They mediate cutaneous recruitment of human CD8 + T cells 16 , 17 . The expression of CXCR6 has been observed in Th17 cells and Tc17 with increased levels of CXCL16 18 . Furthermore, CXCL16 has been demonstrated to regulate the migration of CXCR6-expressing CD8 + T cells and promote Tc17 differentiation 19 , 20 . Additionally, CXCL16 is secreted by macrophages and dendritic cells (DCs) to attract memory T cells 21 . Molecular studies demonstrated that fibrosus cells exposed to IL-1ß, exhibited a notable increase in CXCL16 expression compared to control cells 22 . The precise function of CXCL16 in dendritic cells (DCs) and CXCR6 in γδT and CD8 + T cells, particularly within the context of the epidermal psoriatic milieu, remains to be elucidated. In this study, we demonstrated an epidermal Ube2l3 conditional knockout mouse model exhibited a complete mimicry of the human psoriatic phenotype. This was achieved through the analysis of using bulk RNA levels, protein levels and single-cell RNA sequencing (scRNA-seq). In conclusion, UBE2L3 functions as a keratinocyte-intrinsic suppressor of epidermal IL-17 production in mouse Vγ2 + γδT and human CD8 + T cells via the CXCL16/CXCR6 signaling pathway in psoriasis. RESULTS Cutaneous phenotypes of epidermal deficiency of Ube2l3 mouse models exhibit a high degree of similarity to that observed in human psoriasis Previously, our research demonstrated that in patients with psoriasis, UBE2L3 is predominantly downregulated in the epidermis, with the exception of the dermis 23 . To gain further insight into the underlying mechanisms, we established a Ube2l3 epidermal deficiency mouse model ( Ube2l3 △Epi , Fig. 1 a). The cutaneous phenotype of Ube2l3 △Epi murine models exhibited notable erythema, thickness, and scales as the induction time by tamoxifen (TAM) was prolonged (Fig. 1 b-d). The substantial downregulation of Ube2l3 in the epidermis of Ube2l3 △Epi group was substantiated through the implementation of western blot, bulk RNA sequencing, immunofluorescence and immunohistochemistry staining (Fig. 1 e, 1 f, Fig. S1 a). A typical psoriasis-like skin phenotype was established in back, ear and tail (Fig. 1 g), measured by significantly increased epidermal thickness in Ube2l3 △Epi mice (Fig. 1 h, 1 i, and 1 j), comparable to human psoriatic skin lesions (Fig. 1 k). The increased angiogenesis (CD31), proliferation (Ki67) and immunocytes infiltration (CD3, CD11c), which corresponded to the psoriasis-like phenotype and inflammatory stage (Fig. 1 l, Fig. S1 b-d), thus provide a new insight into the ability to mimic psoriasis disease in the mouse model. A comparative analysis of RNA and protein levels between Ube2l3 △Epi mice and human psoriasis skin To investigate the molecular mechanisms involved in sustaining skin inflammation in the epidermis or dermis, we conducted ex vivo bulk-RNA sequencing and Illumina array analysis of epidermis and dermis respectively from Ube2l3 △Epi mice and control (Ctrl) mice. The principal component analysis (PCA) demonstrated a distinct separation of the four groups (Fig. 2 a). The number of genes in the dermis is significantly lower than in the epidermis when using the same p-value threshold of less than 0.05 and an absolute log2 fold change greater than 1.5 are employed (Fig. 2 b, 2 c, and 2 d). The data were obtained from GSE166388, GSE68937 and GSE68923, and a comparison was made between human psoriatic lesional skin and normal skin from controls in the epidermis (Fig. S2a-b). The genes that were found to be upregulated were collated and analysed by using the Metascape website 24 in order to identify all molecules and enrichment pathways. The Ube2l3 △Epi mouse was found to be closely associated with human psoriasis in the epidermis, specifically in relation to the immune response (Fig. 2 e-f). The enrichment analyses of the dermis, conducted using differentially expressed genes (DEGs), demonstrated a lesser degree of correlation with psoriasis compared to the epidermis (Fig.S2c-d). The genes found to be related to psoriasis belong to the majority of members of the cytokine and chemokine families, including interleukin-1 (IL-1), IL-17, IL-6/IL-23 25 , chemokines and their receptors, antimicrobial peptides (AMPs), tumour necrosis factor (TNF), toll-like receptors (TLRs), as well as representative Type I interferon receptors (IFNRs). The heatmap indicated that these families were all significantly upregulated in the epidermis of Ube2l3 △Epi mice, which is analogous to human psoriasis (Fig. 2 g, 2 h). The Illumina array analysis revealed increase in protein levels of AMPs (S100A8, S100A9), vascular endothelial growth factor (VEGF), chemokines (CCL2, CCL22, CXCL2), and cytokine (IL-17A) in the epidermis of the Ube2l3 △Epi mouse (Fig. 2 i). Furthermore, the iTRAQ analysis revealed an increase in antimicrobial peptides (AMPs), while the Illumina array analysis indicated a tendency towards an increase in tumour necrosis factor (TNF), interleukin 12/interleukin 23p40 (IL-12/IL23 P40), and IL-17A in the human dermis (Fig. 2 j). However, no significant differences were observed in psoriasis-related families in the dermis as that in the epidermis, as indicated by both the enrichment analysis and the heatmap (Fig. S2c-d). Consequently, a comparison of RNA and protein levels between the Ube2l3 △Epi mice and human psoriasis skin revealed a striking similarity. Mouse versus human epidermal lymphocytes subpopulations with respect to γδT and CD8 + T cells secreting IL-17A We next sought to compare subpopulations within the T lymphocytes compartments from human psoriasis skin lesion and Ube2l3 △Epi mouse model. IL-17A, the principal downstream cytokine of IL-23, is most strongly implicated in the pathogenesis of psoriasis, with T lymphocytes secreting it in the greatest quantity 26 . scRNA-seq data showed a prevalence of infiltrating IL-17A secreting γδT (gdT17) cells in the epidermis of Ube2l3 △Epi mice (UE), whereas dendritic epidermal t cells (DETC) were dominant in the epidermis of control mice (CE) (Fig. 3 a, 3 b). The gdT17 emonstrated a high level of expression of Il17a, Il17f and Tcrg-V4 , whereas the DETC displayed a high relative expression of Cd3e, Trdv4, Tcrg-v5 and Fcer1g 27 . Both cell types displayed a lack of Cd4/Cd8a expression (Fig. 3 c). Furthermore, Immune CellAI-mouse 28 was employed to elucidate the infiltration score in various cell types in both the epidermis and dermis. The findings revealed that the infiltration of γδT was markedly more pronounced in the epidermis of Ube2l3 △Epi mice in comparison to the dermis (Fig.S3a). In the historical context, IL-17-producing T cells have been predominantly regarded as CD4 + T cells. However, recent observations have provided growing evidence for the presence of IL-17 producing CD8 + T cells in psoriasis 29 . A similar isolation of human lymphocytes yielded 15 clusters, including IL-17 signalling-related CD8 + T cells (CCL20_Tc17, IL-17A_Tc17, IL-17F_Tc17 and IL26_Tc17), based on expression of CD4 , CD8A , or CD8B ( Fig. 3 d, 3 e, 3 f ) , as observed in other recent studies 30 . Moreover, Th17/Tc17 cytokines, including IL17A , IL17F , IL26 31 ,CCL20 and CXCL13 32 were also employed (Fig. 3 d- 3 f, and Fig.S3b). Another group was defined as cytotoxic T cells (Tc), which were identified by the presence of GNLY , GZMH , HSPA1B and IFNG (Fig. 3 d-f). The simultaneous up-regulation of the T cell receptor signalling pathway, Th17 cell differentiation and Th1 and Th2 cell differentiation was observed in both the mouse gdT17 and human Tc17 clusters (Fig. 3 g, h). The nuclear receptor retinoid-related orphan receptor gamma t ( RORγt, RORC ) has been identified as a pivotal regulator of the differentiation and expansion of Th17 cells 14 . One of the most significant findings was that Rorc, a transcription factor (TF) with a pivotal role in the differentiation and expansion of Th17 cells, was identified as one of the top TFs in the mouse gdT17 cluster (Fig. 3 i). This observation aligns with the characteristics of human psoriasis type 17 cell differentiation 33 . We proceeded to perform flow cytometry on IL-17A source cells derived from Ube2l3 △Epi mice skin lesions and psoriasis patients skin biopsies, in comparison with those from site-matched control mice (Ctrl)/ Ube2l3 △Epi mice. An increase in IL-17A-producing γδT cells was observed in the epidermal samples of Ube2l3 △Epi mice (Epi- Ube2l3 △Epi mice samples) in comparison with those of control mice (Epi-Ctrl). This was in contrast to the observed increase in TNFα secretion (Fig. 3 j, Fig.S3c). In addition, an increase in IL-17A-producing CD8 + T cells (Tc17) was observed in epidermal psoriatic lesions (PSO-Epi) (Fig. 3 k). Following dermal conversion, no notable discrepancy in IL-17A secretion was discerned between the Ube2l3 △Epi mice samples (Der- Ube2l3 △Epi ) and the control mice (Der-Ctrl) (Fig.S3d). The administration of an antibody that neutralises IL-17A to Ube2l3 △Epi mice during the effector phase resulted in a reduction in epidermal thickness, a decrease in the infiltration of lymphocytes (CD3), and an enhanced tendency for epidermal keratinocytes to differentiate (Loricrin) (Fig. 3 l, 3 m and 3 n). Therefore, the Ube2l3 conditional deficiency model of psoriasis in epidermal γδT cells is largely analogous to the IL-17A-secreting CD8 + T cells observed in human psoriasis. Crosstalk with CXCL16-CXCR6 signaling informed ligand receptor analysis The maintenance of homeostasis requires intercellular communication between diverse cell types. An approach to assess such communication is provided by single-cell integration. The initial step involved the tabulation of inferred ligand-receptor interactions across keratinocytes, myeloid cells (myeloids) and NK/T cells (Fig. 4 a, 4 b, 4 c). Subsequently, the most highly expressed ligand-receptor pairs from known major pathways operating in mouse epidermal skin were curated. Within the epidermis, one of the most prominent pairs of NK/T cells demonstrated crosstalk with myeloid cells or keratinocytes through Cxcl16-Cxcr6 signalling (Fig. 4 d-e). Cxcr6 was previously demonstrated to be expressed in T cells, which were predominantly enriched in gdT17 clusters with high expression of Vγ2 and Vγ4 specified markers simultaneously ( Tcrg-V4, 5830411N06Rik and Cd163l ) 34 , 35 (Fig. 4 f). A greater proportion of IL-17A and CXCR6 was observed to be co-expressed in Epi- Ube2l3 △Epi , as evidenced by flow cytometry analysis (Fig. 4 g). The percentage of Vγ2 + γδT cells within the γδT cell population was found to be significantly increased, while the frequency of Vγ3 + cells was observed to be abnormally decreased (Fig. 4 h). The observed up-regulation of IL-17A + Vγ2 + γδT cells indicates that the increase in IL-17A proportion in T cells is driven by Vγ2 + γδT upregulation (Fig. 4 i). The intraperitoneal neutralisation of Vγ2 in Ube2l3 △Epi psoriasis-like mice has led to the conclusion that Vγ2 + γδT may play an important role in this psoriasis-like mouse model. Furthermore, a decreased trend towards psoriasis-like lesions, epidermal thickness (Fig. 4 j-k) and IL-17A proportion of γδT in the epidermis of the anti-Vγ2 group (Fig. 4 l, Fig. S4a) was observed. No significant difference was observed between the dermal IL-17A and TNFα proportions in the isotype and anti-Vγ2 groups (Fig.S4b). In addition, the knock-out of Rag1 in Ube2l3 △Epi mice ( Ube2l3 △Epi -Rag1 −/− ) also exhibited a decreased epidermal thickness compared Ube2l3 △Epi mice (Fig. S4c). These all indicate that Vγ2 + γδT-secreting IL-17A played a pivotal role in the inflammatory environment of the mouse epidermis. We next ascertain the relevance of human psoriasis in epidermal CXCR6 + T cells. The expression of CXCR6 found to be elevated in psoriatic lesional skin at the baseline stage (LS-baseline) in comparison to non-lesional skin (NL) and following the administration of biologic treatment (LS-treated) (Fig. 4 m). A comparison was made between lymphocytes isolated from both adult epidermal healthy skin and epidermal psoriasis samples from the scRNA-seq dataset. The spatial transcriptomics, dotplot analysis and heatmap in the scRNA-seq analysis revealed an abundance of CXCR6 enrichment in the Tc17 population in the epidermis (Fig. 4 n-o, Fig.S4d-e), which is consistent with the immunofluorescence staining and flow cytometry analysis in the epidermis in human psoriasis (Fig. 4 p-q). In conclusion, epidermal IL-17A signalling was demonstrated to be regulated by CXCR6 + Vγ2 + γδ T cells in the mouse model and by CXCR6 + CD8 + T cells in the human subject. IL-17A and IL-1β stimulated secretion of CXCL16 in keratinocytes through STAT3 signaling The scRNA-seq analyses of the Ube2l3 △Epi psoriasis-like mouse model provided evidence to support the hypothesis that keratinocyte subpopulation dysfunction may play a role in the pathogenesis of psoriasis. The proportion of basal and spinous cells between healthy and psoriatic skin was analysed, revealing an increase in both proliferating basal cells (prolif.Basal), specified by Top2a, Mki67 36 , and cycling spinous cells (Spinous II) in the Ube2l3 △Epi mouse epidermis (Mouse-UE) compared to the control epidermis (Mouse-CE). This suggests that the basal layer exhibited hyperproliferation (Fig. 5 a-b). We therefore conducted a detailed investigation into the up-regulated expression of CXCL16 in Mouse-UE, which was predominantly expressed in the prolif.Basal cluster (Fig. 5 d-e). This finding was consistent with the CXCL16 expression observed in CD45- cells, as analysed by flow cytometry (Fig. 5 f). We therefore proceeded to investigate the relationship between Ube2l3 deficiency and CXCL16 secretion. Previous studies have demonstrated that the overexpression of UBE2L3 reduces its binding to TRIM21, consequently leading to a decrease in STAT3 pathway activity and a reduction in the level of the IL-1β precursor (pro-IL-1β). Furthermore, molecular studies demonstrated that annulus fibrosus cells exposed to IL-1β, but not TNF-α, exhibited a notable elevation in CXCL16 expression relative to control cells 22 . The results demonstrated that both CXCL16 and IL-1β were elevated in Ube2l3 -deficient keratinocytes (KC- Ube2l3 △) relative to control cells (KC-Ctrl) (Fig. 5 g-i). Upon stimulation of keratinocytes with or without recombinant mouse IL-17A (rmIL-17A) and recombinant IL-1β (rmIL-1β), the protein level of CXCL16 was markedly increased, exhibiting a similar trend to that of STAT3 signalling (Fig. 5 k). Furthermore, CXCL16 was found to be upregulated in epidermal Ube2l3 △Epi mice ( Ube2l3 △Epi− Epi) (Fig. 5 l). In human keratinocytes stimulated with IL-17A, an increase in CXCL16 mRNA levels was observed (Fig. 5 m). Similarly, the same up-regulated tendency of CXCL16 in psoriasis in epidermis (PSO-Epi) was observed in CD45- cells by flow cytometry (Fig. 5 n-o). Upon stimulation of normal human epidermal keratinocytes (NHEKs) with IL-1β and IL-17A, a significant up-regulation of CXCL16 was observed in lysates, accompanied by activation of the STAT3 pathway (Fig. 5 p). In conclusion, the data collectively indicate that CXCL16 is secreted by Ube2l3 -deficient keratinocytes and stimulated by IL-1β and IL-17A. This suggests that CXCL16 may act as a chemoattractant for T cells, which in turn secrete IL-17A, forming a positive feedback loop in psoriatic keratinocytes. Up-regulated CXCL16 secreted by cDC2/mDC controlling CXCR6 γδT/CD8T IL-17A secretion in psoriatic epidermis scRNA-seq and bulk RNA sequencing analysis indicated the potential involvement of CXCL16-CXCR6 signalling between myeloid cells and natural killer (NK)/T cells in the epidermis of Ube2l3 △Epi psoriasis-like mice, with no evident impact on the dermis (Fig. 4 d-e, Fig.S5a-c). Therefore, we proceeded to evaluate the proportion of CXCL16 in myeloid cells, identifying a notable presence in the migrating dendritic cell (mDC) and cDC2 (type 2 DC) population, specifically marked by Itgam, Irf4, Fscn1, Cacnb3 and Ccr7 37 (Fig. 6 a-c, Fig.S5e). The results of the pseudo-time analysis indicated that Cxcl16 was predominantly expressed in mDC_cDC2 and Mac (macrophage) (Fig. 6 d). The same trend was corroborated by flow cytometry, which demonstrated that cDC2 was the predominant cell type secreting CXCL16 in the epidermis (Fig. 6 e). Following the knockout of the Zbtb gene in Ube2l3 △Epi mice, Ube2l3 △Epi -Zbtb −/− mice were generated after TAM induction (Fig. 6 e). The epidermal thickness of the psoriasis-like lesions was found to be decreased in conjunction with the down-regulation of CD11c and keratin 14 (K14) (Fig. 6 f-g). Furthermore, we observed a notable reduction in the proportion of γδT cells secreting IL-17A in Ube2l3 △Epi -Zbtb −/− (Fig. 6 h). In a human myeloid single-cell analysis, a comparable population of mDC and cDC2 was identified based on the expression of CCR7, LAMP3, CLEC10A, CD1C 38 , 39 (Fig. 6 j, Fig.S5f-g), The results demonstrated that CXCL16 was markedly elevated in the epidermis of PSO (Epi-PSO) specimens, as evidenced by heatmap analysis and flow cytometry (Fig. 6 i-k). Similarly, a comparable trend is observed in myeloid cells, with the involvement of analogous pathways, such as the regulation of cell activation (Fig.S5h-i). Therefore, CXCL16 was released by the cDC2/mDC population simultaneously and may be involved in the regulation of γδT17 secretion in the psoriatic epidermis. CXCL16 facilitates Vγ2 γδT/ CD8T migration, proliferation and IL-17A release in psoriatic epidermis In wild-type mice, Vγ2 + γδT was observed to be absent in the epidermis and present in the dermis 40 . In psoriasis-like Ube2l3 △Epi mice, the percentage of Vγ2 + γδT cells was significantly increased in the dermis (Fig. 7 a). Epithelial cells lining the mucosal surfaces play a crucial role in this process by producing specific chemokines, such as CXCL16, which serve as attractants for γδT cells expressing the corresponding chemokine receptors 41 . Furthermore, IL-23R signalling is responsible for the generation of CXCR6 + cells from IL-17 + SLAMF6 + cells 42 . It was therefore postulated that there is a relationship between CXCL16 and CXCR6 + Tc17/γδT17. The migration assay demonstrated that CXCL16 exerted a migratory effect on dermal Vγ2 + γδT (Fig. 7 b-c). In accordance with the limited research on the conversion of DETC to Vγ2 + γδT17, we conducted a pseudotime analysis that revealed the non-near differentiation of Vγ2 + γδT17 and DETC (Fig. 7 d). Next, epidermal and dermal Vγ2 + γδT cells were sorted and stimulated with Cxcl16 (Fig. 7 e). The results demonstrated that the percentage of Ki-67 and IL-17A was increased in two-week-old TAM-induced Ube2l3 △Epi mice (Fig. 7 f). Furthermore, the secretion of IL-17A was increased in Vγ2 + γδT cells in 6 week-TAM induced Ube2l3 △Epi mice (Fig. 7 g). CXCL16 neutralization in Ube2l3 △Epi mice resulted in a reduction in the epidermal thickness of psoriasis-like lesions (Fig. 7 i). Upon stimulation of the human epidermal psoriasis epidermis (Human-Epi-CD3 + T) with recombinant human CXCL16 (rhCXCL16), an increase in the percentage of IL-17A secretion was observed in CD3 + T cells, particularly in CD8 + T cells (Fig. 7 j-k). In wild-type (WT) mice, it was observed that recombinant mouse CXCL16 (rmCXCL16) injection resulted in an increase in ear epidermal thickness, in comparison to rmIL-23 injection in the right ear (Fig. 7 l, 7 m and 7 n). Therefore, CXCL16 can facilitate the migration, proliferation and IL-17A secretion of Vγ2 + γδT cells in mouse epidermis and CD8 + T cells in human psoriatic epidermis. DISCUSSION In this study, scRNA-seq from human psoriasis skin was integrated to map subpopulation composition and intercellular communication within the major compartments of the skin (keratinocytes, myeloid cells and NK/T cells), thereby generating a comprehensive resource for hypotheses regarding the epidermal immune environment in psoriasis. A psoriasis-like lesion mouse model was generated, which may facilitate further insights into the pathogenesis of psoriasis 3 . The data were compared with mouse skin scRNA-seq data to facilitate cross-species comparisons of differentiation dynamics and ligand-receptor pathways. In particular, the CXCL16-CXCR6 signalling pathway was identified as a highly active cluster in keratinocytes/myeloids and NK/T cells (Graphical abstract). Some research has indicated that pathogenic CXCR6 + Th17 populations are induced in autoimmunity 42 . In the context of inflammatory diseases, the activation of CXCR6 + T cells has been identified as a key factor in the differentiation of T cells into Th17 and the subsequent secretion of pro-inflammatory cytokines 43 . In human, CXCR6-expressing cells were absent in the epidermis of healthy skin 44 . In contrast, CXCR6 + CD8 + T cells were significantly increased in both peripheral blood and skin in psoriasis patients 43 . In mouse model, previous sltudies have demonstrated that dermal γδT cells are the primary IL-17-producing cells in the skin that respond to IL-23 stimulation in an IMQ (imiquimod) induced psoriasis-like mouse model 45 . However, there is a paucity of research examining the epidermal CXCL16-CXCR6/IL-17 signalling pathway in psoriasis patients and psoriasis-like mouse models. In addition to it, IL-17-induced effector cells in skin inflammation are keratinocytes 46 . In our studies, we found that CXCL16 was secreted by Ube2l3 -deficient keratinocytes, stimulated by IL-1β through the STAT3 pathway, together with CXCL16 from cDC2 and mDC, promoted the migration and proliferation of γδT17 cells in the epidermis of Ube2l3 △Epi mice. The same activated pathway was found in human psoriasis epidermis and the key cell was CD8 + Tc17 (Fig. 3 ). We first compared the psoriasis-like mouse model and human psoriasis patients in the epidermis and found both CXCL16-CXCR6 signaling played a crucial role in psoriatic cell communication. Our single cell trajectory analysis shed the light on the root of convert and demonstrated the reduced potential of DETC to Vγ2 + γδT17 (Fig. 7 d). The increased epidermal Vγ2 + γδT17 was derived from the dermis and CXCL16 from keratinocytes/dendritic cells may further facilitate Vγ2 + γδT17 proliferation and differentiation (Fig. 7 f-g). We also found that cutaneous injection of CXCL16 neutralising antibody contributed to the amelioration of psoriasis-like lesion in Ube2l3 △Epi mice (Fig. 7 i). Therefore, CXCL16 may be another therapeutic target and be suitable for use in a precision medicine approach to psoriasis treatment, especially in patients with elevated CXCL16. Overall, our findings indicate that CXCR6 + γδT17/ Tc17cells are prevalent within the epidermal immune microenvironment of psoriasis 8 , 47 . The elevated level of CXCL16 produced by Ube2l3 -deficient serves to augment the differentiation and recruitment of γδT17/ Tc17cells, thereby establishing a positive feedback loop that facilitates keratinocyte advancement in psoriasis. Furthermore, we demonstrated that the deletion of CXCL16 inhibits the progression of psoriasis-like lesions in Ube2l3 △Epi mice and exhibits a therapeutic effect in vivo. The findings of our study offer valuable insights that may inform the development of a therapeutic approach utilising anti-CXCL16 neutralising antibodies, with the potential to provide alleviation of psoriasis. METHODS Mice and ethics All participating patients with plaque psoriasis patients and healthy donors gave written informed consent before inclusion. A skin biopsy with a diameter of 1–2 cm was taken from healthy donors’ or psoriasis patients’ extremities and/or back. The study was approved by the Ethics Committee of the Second Affiliated Hospital, Zhejiang University School of Medicine (2020-No.135). In the animal study, C57BL/6 mice was supplied by SLAC Laboratory Animal Co. (Shanghai, China), Zbtb46-DTR mice (Jackson Laboratories), Ube2l3 fl/fl (Cyagen)and were housed under specific pathogen-free (SPF) conditions. Ube2l3 fl/fl (mice were crossed to K14-Cre ERT mice (Jackson Laboratories) to generate Ube2l3 epidermis-specific knockout mice ( Ube2l3 fl/fl - K14-Cre ERT ). When Ube2l3 fl/fl - K14-Cre ER was induced by tamoxifen for 6 weeks, we called them Ube2l3 △Epi mice. Ube2l3 fl/fl - K14 - Cre ERT were crossed to Zbtb46-DTR mice to generate DC knockout in Ube2l3 △Epi mice ( Ube2l3 △Epi - Zbtb −/− ). Ube2l3 fl/fl - K14 - Cre ERT were crossed to Rag1 −/− mice (GemPharmatech) to generate DC knockout in Ube2l3 △Epi mice ( Ube2l3 △Epi - Rag1 −/− ). Sex- and age-matched animals between 12 and 15 weeks of age were used for experiments. Preliminary experiments were performed to determine proper sample size. All animal experiments were performed in accordance with protocols approved by the Second Affiliated Hospital, Zhejiang University School of Medicine Animal Care Committee (2019-No.072). Flow cytometric analysis of skin tissue-derived cells Mouse/human skin biopsy tissue was immersed in 0.5% dispase (GibcoTM, USA) at 4°C overnight in order to separate the epidermis and dermis. The epidermis was digested with 0.25% trypsin (Thermo Fisher Scientific, USA), while the dermis was digested with 1 mg/ml collagenase (Sigma-Aldrich, USA). After neutralization with serum, the mixture was centrifuged at 1000 rpm for 5 min, and the cells were resuspended in PBS to obtain a single-cell suspension. Epidermal and dermal single-cell suspensions were prepared as described above. The cells were first stimulated with Cell Activation Cocktail (with Brefeldin A, BioLegend) together with 2 µg/ml resiquimod (Sigma) for 6 hours. After stimulation, the cells were first stained with a Zombie UV Fixable Viability Kit (BioLegend) for 15 minutes and then incubated with the Fc receptor blocker TruStain FcX (BioLegend) for 10 minutes. Then, the cells were incubated with mixed cell surface antibodies for 30 minutes at 4°C in the dark. For intracellular staining, cells were fixed and permeabilized by using the BD Cytofix/Cytoperm Kit (BD Biosciences) or Foxp3/Transcription Factor Staining Buffer Set (eBioscience) and then incubated with anti-cytokine antibodies for 30 minutes at 4°C in the dark. The antibodies used were purchased from BioLegend, BD Biosciences, or eBioscience, as listed in supplementary methods. The data were acquired by CytoFLEX LX (Beckman Coulter) and analyzed by CytExpert Software (Beckman Coulter). Single-cell RNA sequencing data analysis Processed human single-cell RNA-sequencing (scRNA-seq) datasets from our datasets, sequenced by performed by PLTTech Inc. (Hangzhou, China). For human skin integration of the psoriatic epidermis (PE) and normal epidermis (NE) was analyzed with the Seurat R package (versions 3.0.0 and 4.0.3) 48 . Cells with > 20% mitochondrial gene percentage, minimum of 350 genes and maximum of 4500 detected were filtered from downstream analyses. Samples were split by donor identity (referred to as sample in metadata in analysis scripts and Seurat objects) into individual objects by donor, which were each normalized and processed with the NormalizeData and FindVariableFeatures functions with default parameters. Integration anchors were determined with these donor objects using the first 30 dimensions and integrated using the IntegrateData function on the first 30 dimensions. Using this integrated object, dimensionality reduction was carried out after ScaleData with principal component analysis and Uniform manifold approximation and projection (UMAP) on the first 30 principal components to generate UMAP. T lymphocytes (Human NE, Human PE) clustering was performed at resolution 0.2 after FindNeighbors was run on the first 30 dimensions. Myeloid cells clustering was performed at resolution 0.5 after FnidNeighbors was run on the first 30 dimensions. Similarly, for mouse skin integrated data, a Seurat object was created with 10x cellranger outputs. SingleCell Data Analysis was performed using the OmicStudio tools created by LC-BIO Co., Ltd (HangZhou, China) at https://www.omicstudio.cn/cell , and was analyzed with the Seurat R package (4.1.0, R version 4.1.3). Anti-IL-17A/ Vγ2/ CXCL16 treatment The procedure of anti-IL-17A treatment was attached in Fig. 3 l with 50ug/g/week hamster anti-mouse mAb (17F3) by subcutaneous injection (s.c.) and respective hamster IgG isotype control antibody (BE0091, both Bio X Cell) (Fig. 4 j). Anti-Vγ2 treatment was performed by intraperitoneal injection (i.p.) of the mice with 10 µg/g/week hamster anti-mouse mAb (UC3-10A6) and respective hamster IgG isotype control antibody (BE0091, both Bio X Cell) (Fig. 4 j). Anti-mouse CXCL16 treatment (R&D, AF503), was performed by subcutaneous injecti4on (s.c.) of the mice with 4ug/g/week and respective hamster IgG isotype control antibody (Fig. 7 h). Primary keratinocyte culture The separated epidermis was digested with 0.25% trypsin (Thermo Fisher Scientific, USA). The digested epidermal keratinocytes were neutralized with FBS (Gibco, USA) and centrifuged at 1000 rpm. Cells were cultured to 70% confluence in EpiGRO™ Human Epidermal Keratinocyte Complete Culture Media Kit (Millipore, USA, SCMK001) in a humidified incubator with 5% CO 2 at 37°C. Recombinant mouse IL-23/CXCL16 treatment Mice were injected intradermally (i.d.) with 20 µL PBS/0.1% bovine serum albumin (BSA) containing 0.5 µg recombinant mouse (rm)IL-23 or recombinant mouse (rm)CXCL16 in the right ear using a 29-gauge needle, every other day for 14 days. Quantification and statistical analysis Statistical analyses were performed with GraphPad Prism, versions 9–10 (GraphPad Software, San Diego, CA), or R, versions 3.5.1, 3.61, and 4.0.2. Parameters such as number of replicates; the number of independent experiments; measures of center, dispersion, and precision (mean SD or SEM); statistical test; and significance are reported in figures and figure legends. P values are indicated as follows: ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001. Declarations Funding This study was supported by grants from the National Natural Science Foundation of China (No. 82230104, 81930089, 81773318, 82404117, 82103709 and 82303999). Author Contributions Conceptualization: XYC, LRY; Formal Analysis: XYC, XYM; Funding Acquisition: XYM; Investigation: XYC, LRY, XYM, NCF, XBC; Methodology: BXY, NCF, YZC, XYM, XYC, SQC; YXZ, MYL; Project Administration: XYM; Resources: XYM; Supervision: MZ, XYM; Validation: XYC, YZ, NCF; Writing and original Draft Preparation: XYC, LRY; Writing - Review and Editing: LRY, XYC, XYM. Competing interests The authors declare no competing interests. Data and materials availability: All data needed to evaluate the conclusions in the paper are present in the paper or the Supplementary Materials. The bulk-RNA-seq data and scRNA-seq data are available in the Gene Expression Omnibus (GEO) repository database and are submitting. The Ube2l3 fl/fl mice developed for this study will be shared upon request. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-5973089","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":421196307,"identity":"a530b635-ea44-4628-92d0-e16a0281ef5d","order_by":0,"name":"Xiao-Yong 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\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003efl/fl\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e-K14\u003c/em\u003e\u003csup\u003e\u003cem\u003ecreERT \u003c/em\u003e\u003c/sup\u003emice by oral gavage of TAM (200mg/kg) for 6 weeks. (b) The gradual change of phenotype of \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi \u003c/sup\u003emice in TAM induction in week 2, 4 and 6. (c-d) Incidence rate of psoriasis-like phenotype in the back and ear of \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi \u003c/sup\u003emice during 6-week TAM induction. The average change of PASI in the back of \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi \u003c/sup\u003emice. (e)The western blot results from epidermis between \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi \u003c/sup\u003emice group and Ctrl group in epidermis. Image J analysis of western blot of Ctrl group(n=6) and \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e\u003csup\u003e \u003c/sup\u003egroup(n=4), **p\u0026lt;0.01. (f) FPKM of \u003cem\u003eUbe2l3\u003c/em\u003e between \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi \u003c/sup\u003emice group and Ctrl group in epidermis by using RNA-seq analysis, ****p\u0026lt;0.0001. (g) Changes of phenotypes and skin thickness of Ctrl and \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003emice mouse ear, skin and nail after tamoxifen application, **p\u0026lt;0.01, *p\u0026lt;0.05. (h) Changes of back skin and ear thickness in mouse skin after tamoxifen application (Ctrl, n=7;\u003cem\u003e Ube2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e-skin, n=5). (i) H\u0026amp;E staining to compare histopathological differences between Ctrl and \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e\u003csup\u003e \u003c/sup\u003emice after applying tamoxifen (scale bar is 50μm inback skin, ear skin, scale bar is 250μm in tail skin). (j)The statistics analysis of epidermal thickness of back skin and ears from measuring H\u0026amp;E staning (μm). ***p\u0026lt;0.001, **p\u0026lt;0.01. (k) H\u0026amp;E staining of human skin between healthy donors (HD) and psoriatic lesions (PSO). (l) Statistics of immunofluoresence of vascluar genesis(CD31\u003csup\u003e+\u003c/sup\u003e), T cells (CD3\u003csup\u003e+\u003c/sup\u003e) and immunohistology of proliferation(Ki-67) respectively in the back skin of\u003cem\u003e \u003c/em\u003eCtrl and \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e\u003csup\u003e \u003c/sup\u003emice (scale bar=100μm). **p\u0026lt;0.01.\u003c/p\u003e\n\u003cp\u003eTAM, tamoxifen. H\u0026amp;E, hematoxylin-eosin.\u003c/p\u003e","description":"","filename":"Picture1.png","url":"https://assets-eu.researchsquare.com/files/rs-5973089/v1/610552ebda63492e0210d9dc.png"},{"id":77427846,"identity":"2765f66f-3d97-48aa-a028-c698d7152a84","added_by":"auto","created_at":"2025-02-28 13:31:55","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":3381556,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSimilarity between epidermal deficiency of \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eUbe2l3\u003c/strong\u003e\u003c/em\u003e\u003csup\u003e\u003cstrong\u003e△Epi \u003c/strong\u003e\u003c/sup\u003e\u003cstrong\u003emice and human psoriasis in bulk-RNA and protein levels.\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e(a) Principal component (PCA) analysis (U_E, n = 6, U_D, n = 6, C_E, n = 8, C_D, n = 8) was performed on all samples of \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e mice and Ctrl mice of epidermis and dermis. U_E, epidermis of \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e \u003c/em\u003emice, U_D, dermis of \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e \u003c/em\u003emice, C_E, epidermis of control mice, C_D, dermis of control mice. (b) Volcano plot of differentially expressed genes between the epidermis of \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e \u003c/em\u003emice and Ctrl mice. The red (1631) on the right is the up-regulated gene of the epidermis of \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi \u003c/em\u003e\u003c/sup\u003emice relative to the epidermis of Ctrl mice, and the green (592) on the left is the down-regulated gene. p-val\u0026lt;0.05 and |log2foldchange|\u0026gt;1.5. (c) The difference gene number between the dermis of \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e \u003c/em\u003emice and Ctrl mice was analyzed. The number of up-regulated gene is 288, the number of down-regulated gene is 245. (d) The number of differential genes in all samples of epidermis and dermis of the two groups were compared. (e) For all up-regulated genes in \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e mice, the results of metascape enrichment analysis showed a comprehensive ranking of enrichment analysis results. The darker the color is and the greater the - log10 (P), the more obvious the difference is and the higher the enrichment fraction is. (f) The metascape analysis of human up-regulated gene in GSE166388, GSE68937 and GSE68923. (g-H) Heatmap in FPKM expression of representative cytokine and chemokine families of IL-1, IL-17, IL-6/IL-23, AMPs, TNF, TLRs, as well as IFNRs in the epidermis of mouse(f) and human groups (g). (i) Significant protein level of S100A8, S100A9, VEGF, CCL2, CCL22, CXCL2 in Ilumina array analysis in epidermal Ctrl mice (Epi-Ctrl) and \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e \u003c/em\u003emice (Epi-\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e)\u003c/em\u003e. Epi-Ctrl, n=5, Epi-\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e, n=4 or 5. Elisa results of Epi-Ctrl and Epi-\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e. Epi-Ctrl, n=3, Epi-\u003cem\u003e Ube2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e, n=2. ***p\u0026lt;0.001, **p\u0026lt;0.01, *p\u0026lt;0.05. (j) iTRAQ results of S100A8 and S100A9 in epidermal healthy donor skin (Epi-HD) and psoriasis lesion (Epi-PSO). Epi-HD, n=15, Epi-PSO, n=16. IL-17A, TNFa, and IL-12/IL-23p40 illumina array results in epidermal and dermal of healthy donors (Der-HD) and psoriasis patients (Der-pso) respectively. Epi-HD, n=3 or 4, Epi-PSO, n=3 or 4. Der-HD, n=4, Der-PSO, n=4. **p\u0026lt;0.01, *p\u0026lt;0.05, ns=not significant.\u003c/p\u003e","description":"","filename":"Picture2.png","url":"https://assets-eu.researchsquare.com/files/rs-5973089/v1/381e0814da3d5e1072819c1b.png"},{"id":77427852,"identity":"7caa7649-560f-44ff-b50d-3f87421e04b3","added_by":"auto","created_at":"2025-02-28 13:31:55","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":6463480,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eMouse versus human epidermal lymphocytes subpopulation concerning epidermal CD8\u003c/strong\u003e\u003csup\u003e\u003cstrong\u003e+\u003c/strong\u003e\u003c/sup\u003e\u003cstrong\u003e Tc17 and γδT17.\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e(a) UMAP visualization of isolated lymphoid cells in mouse control epidermal skin (Mouse-CE) and \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e epidermal skin (Mouse-UE). CD8T=CD8\u003csup\u003e+\u003c/sup\u003e T cell, DETC=dendritic epidermal T cells, NK=natural killer cell, ILC=innate lymphoid cell, Unknown= unknown cell, Treg=regulatory T cell, gdT17= IL-17A secreting γδT, Cyc-T, cycling T cell, CD4=CD4\u003csup\u003e+\u003c/sup\u003e T cell, each group, n=3. (b) The comparison between the percentage of each cluster in two group (CE and UE). ** p \u0026lt; 0.01, * p \u0026lt; 0.05. (c) Dot plot showing expression of marker genes of each cell cluster found in mouse epidermal skin. (d) Integrated UMAPs of isolated lymphoid cells of human cells labeled by cluster/subpopulation on the right. CCL20_Tc17 = cytotoxic CD8+ T cells expressing CCL20, CXCL13_Th17= T cells expressing CXCL13, FOXP3_Treg=regulatory T cell expressing FOXP3, GNLY_Tc= cytotoxic CD8\u003csup\u003e+\u003c/sup\u003e T cells expressing GNLY. (e) All 15 T cell subsets were concluded as 8 different groups in human normal epidermis (Human-NE) and psoriatic epidermis (Human-PE) by single cell RNA sequencing (scRNA-seq) analysis. (f) Dot plot showing the representative expressed genes in T cell subsets. (g) Using KEGG enrichment to analyze different genes in mouse single cell gdT17 subsets in Mouse CE and Mouse UE two groups. (h) KEGG enrichment to analyze different genes in mouse single cell Tc17 subsets in Human NE and Mouse U_E two groups. Red arrows (g-h) are T cell receptor signaling pathway, Th17 cell differentiation, Th1 and Th2 cell differentiation. (i) Pyscenic analysis of mouse gdT17 cluster and Rorc was one of top increased transcription factors (TFs). (j) Flow cytometry showed the percentage of TNFα and IL17A in the epidermis of control mouse (Epi-Ctrl) and \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e(Epi\u003cem\u003e-Ube2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e). Right bar charts showing the proportions of IL-17A\u003csup\u003e+\u003c/sup\u003e CD45\u003csup\u003e+\u003c/sup\u003e /CD45\u003csup\u003e+\u003c/sup\u003e cells, IL-17A\u003csup\u003e+\u003c/sup\u003eCD3\u003csup\u003e+\u003c/sup\u003e/CD3\u003csup\u003e+\u003c/sup\u003e T cells, IL-17A\u003csup\u003e+\u003c/sup\u003eαβT/αβT cells, IL-17A\u003csup\u003e+\u003c/sup\u003eγδT/γδT cells(**，p\u0026lt;0.01; *，p\u0026lt;0.05, Ctrl, n=5,\u003cem\u003e Ube2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e,\u003c/em\u003e n=5). (k) The flow cytometry results of TNFα and IL17A in the epidermal lymphoid cells of healthy donors (Epi-HD) and psoriatic patients (Epi-PSO). Below bar charts showing the proportions of IL-17A\u003csup\u003e+\u003c/sup\u003e CD3\u003csup\u003e+\u003c/sup\u003e /CD3\u003csup\u003e+\u003c/sup\u003e T cells, TNFα\u003csup\u003e+\u003c/sup\u003e CD3\u003csup\u003e+\u003c/sup\u003e /CD3\u003csup\u003e+\u003c/sup\u003e T, IL-17A\u003csup\u003e+\u003c/sup\u003eCD4\u003csup\u003e+\u003c/sup\u003e/IL-17A\u003csup\u003e+\u003c/sup\u003eCD3\u003csup\u003e+\u003c/sup\u003e T cells, IL-17A\u003csup\u003e+\u003c/sup\u003eCD8\u003csup\u003e+\u003c/sup\u003e/ CD3\u003csup\u003e+\u003c/sup\u003e T cells (**，p\u0026lt;0.01; *，p\u0026lt;0.05, HD, n=5, PSO\u003cem\u003e,\u003c/em\u003e n=12). (l) Schematic strategy for investigating the effects of anti-IL-17A mAb or IgG isotype on tamoxifen induced \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi \u003c/em\u003e\u003c/sup\u003emice through intraperitoneal injection. (m) The phenotype and H\u0026amp;E staining of IgG isotype group and anti-IL-17A mAb group in fourth week. right bar charts showed the significant difference of epidermal thickness ( **，p\u0026lt;0.01). (n) Immunofluorence staining of CD3 and Loricrin in two group.\u003c/p\u003e","description":"","filename":"Picture3.png","url":"https://assets-eu.researchsquare.com/files/rs-5973089/v1/f2b2723b74d6f35fc026cb78.png"},{"id":77427851,"identity":"64d59fbd-8394-4776-805d-05471d1e0e14","added_by":"auto","created_at":"2025-02-28 13:31:55","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":7744184,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eActivated IL-17 signaling was mainly secreted by epidermal CXCR6\u003c/strong\u003e\u003csup\u003e\u003cstrong\u003e+\u003c/strong\u003e\u003c/sup\u003e\u003cstrong\u003e Vγ2\u003c/strong\u003e\u003csup\u003e\u003cstrong\u003e+ \u003c/strong\u003e\u003c/sup\u003e\u003cstrong\u003eγδT in mouse and CXCR6\u003c/strong\u003e\u003csup\u003e\u003cstrong\u003e+ \u003c/strong\u003e\u003c/sup\u003e\u003cstrong\u003eCD8\u003c/strong\u003e\u003csup\u003e\u003cstrong\u003e+ \u003c/strong\u003e\u003c/sup\u003e\u003cstrong\u003eT in human psoriasis.\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e(a) The UMAP visualization shows 6 immune and non-immune cell classes conserved epidermis across UE and CE, delineated by Louvain clustering, profiled 39188 cells. cyc-T, cycling T cell, Krt, keratinocytes, Mel, melanocytes, NK/T, natural kill T cells/ T cells, SG, sebaceous gland. (b) Immune cell population marker transcript expression levels (y axis) for the 6 immune cell and non-immune cell populations (x axis). Size of dots represents the fraction of cells expressing a particular marker, and color intensity indicates mean normalized scaled expression levels. (c) Bar charts showing the proportions of 6 types of cell states in control epidermis (CE) and \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e (UE) groups. (d) Dot plot of top ligands and receptors from 6 subsets predicted to interact with each cluster between CE and UE. Red arrow: Cxcl16-Cxcr6 pairs was significantly up-regulated from myeloid cells into NK/T cells. (e) Dot plot of CXCR6_CXCL16 signaling pathway and other top ligands in 6 immune and non-immune cell classes. (f) Dot plot showing the differentially expressed Vγ2 and Vγ4 specified marker (\u003cem\u003eTcrg-V4, 5830411N06Rik and Cd163l\u003c/em\u003e) in T cell subsets. (g) Represent flow cytometry plots and proportions of IL-17A and CXCR6 in CD3\u003csup\u003e+\u003c/sup\u003eT between Epi-Ctrl and Epi\u003cem\u003e-Ube2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e. The proportion of IL-17A\u003csup\u003e+\u003c/sup\u003eCXCR6\u003csup\u003e+\u003c/sup\u003eT was increased in Epi\u003cem\u003e-Ube2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e. **，p\u0026lt;0.01, Epi-Ctrl, n=6, Epi\u003cem\u003e-Ube2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e, n=6. (h) Representative Flow cytometry plots and quantification of the percentage of Vγ2 and Vγ3 cells in γδT cells in Epi-Ctrl and Epi\u003cem\u003e-Ube2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e groups. *，p\u0026lt;0.05, ****，p\u0026lt;0.0001, Epi-Ctrl, n=5, Epi\u003cem\u003e-Ube2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e, n=6. (i) Flow cytometric compared analysis of the percentage of TNFa\u003csup\u003e+\u003c/sup\u003e and IL-17A\u003csup\u003e+\u003c/sup\u003e Vγ2\u003csup\u003e+\u003c/sup\u003ecells in Vγ2\u003csup\u003e+\u003c/sup\u003e γδT cells between Epi-Ctrl and Epi\u003cem\u003e-Ube2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e groups. ***，p\u0026lt;0.001, Epi-Ctrl, n=5, Epi\u003cem\u003e-Ube2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e, n=6. (j) Schematic experimental design showing \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi \u003c/sup\u003emice treated with or without anti-Vγ2mAb (10ug/g/week) intraperitoneally in four weeks. (k) Representative phenotype of \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi \u003c/sup\u003emice treated with or without anti-Vγ2mAb (Isotype group, Vγ2 antibody group) before (Week 0) and after 4-week treatment (Week 4). Histological analysis of H\u0026amp;E staining was calculated and showed a decrease in Vγ2 antibody group. (**, p\u0026lt;0.01, n=5 per group). Immunofluorence analysis of Vγ2 (red) and CD3 (green) was showed. white triangle, double stained Vγ2 and CD3. (l) Proportion of L-17A\u003csup\u003e+\u003c/sup\u003e γδT cells in γδT in Epi-Ctrl, Epi\u003cem\u003e-Ube2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e and the epidermis of mouse \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e group treated with anti-Vγ2mAb (Epi-\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi-\u003c/sup\u003e Vγ2Ab). ***，p\u0026lt;0.001, **，p\u0026lt;0.01, ns, not significant, Epi-Ctrl, n=5, Epi\u003cem\u003e-Ube2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e, n=6, Epi-\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi-\u003c/sup\u003e Vγ2Ab, n=3). (m) FPKM of CXCR6 in GSE53552 (left), GSE14905, and GSE117239 (right). ****，p\u0026lt;0.0001, **，p\u0026lt;0.01, ns, not significant. (n) UMAP plots of CXCR6 in human lymphoid cells in NE and PE group by scRNA-seq analysis. (o) Heatmap results showed the average expression of CXCR6 and other markers in 8 groups in human lymphoid cells in scRNA-seq analysis. (p) Immunofluorescence staining of psoriatic skin for CD3(green), CXCR6 (red), CD8 (purple), and DAPI (blue). White triangle indicated CXCR6\u003csup\u003e+ \u003c/sup\u003eand CD8\u003csup\u003e+\u003c/sup\u003e T cells. Image is representative of n = 3 donors. Scale bar, 50um. (q) Representative flow cytometric plots and percentage of IL-17A\u003csup\u003e+\u003c/sup\u003eCXCR6\u003csup\u003e+\u003c/sup\u003e T cells in CD3\u003csup\u003e+ \u003c/sup\u003eT cells in Epi-HD and Epi-PSO. The proportion of CD4\u003csup\u003e+\u003c/sup\u003e and CD8\u003csup\u003e+\u003c/sup\u003e in IL-17A\u003csup\u003e+\u003c/sup\u003eCXCR6\u003csup\u003e+\u003c/sup\u003eT cells. **，p\u0026lt;0.01, ns, not significant.\u003c/p\u003e","description":"","filename":"Picture4.png","url":"https://assets-eu.researchsquare.com/files/rs-5973089/v1/87bd64c9cf3a1015b8ade7c9.png"},{"id":77428914,"identity":"0634c1a2-8324-47da-9b98-1f8534325347","added_by":"auto","created_at":"2025-02-28 13:39:55","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":418806,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eIL-17A and IL-1β stimulated secretion of CXCL16 in \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eUbe2l3 \u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003eknock out keratinocytes through STAT3 signaling\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e(a)The single cell profile of Krt (keratinocytes) about nine classes conserved epidermis across UE and CE, delineated by Louvain clustering. Basal, basal keratinocytes, specified by Krt14 and Krt5, spinous cell, specified by Krt1 and Krt10, granular cell, specified by Spink5 and Calm5, Krt17, Krt79, hair follicle stem cells (HFSC) related by Cd34, Postn, Ptn and Fst. (b) Bar charts showing the proportions of different types of keratinocyte cells states in control and \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e skin. (c) Representative markers used in classifying keratinoctyes. Transcriptional expression levels (y axis) for the nine lymphocytes cell populations (x axis). Size of dots represents the fraction of cells expressing a particular marker, and color intensity indicates mean normalized scaled expression levels. (d) Top up-regulated genes in krt subset in scRNA-seq of \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi \u003c/sup\u003emice versus ctrl mice in Krt group. (e) Dot plot of cytokines and chemokines, including Cxcl16, Ccl20, Il23a, Tnf and Il1b compared in each group in Krt subset. red arrow is Prolif.cell group. (f) Flow cytometric plots and percentage of CXCL16 expression in CD45\u003csup\u003e-\u003c/sup\u003e cells in Epi-Ctrl, Epi-IMQ and Epi-\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e group. ***，p\u0026lt;0.001, **，p\u0026lt;0.001, ns, not significant. Epi-Ctrl, n=6, Epi-IMQ,n=6, Epi-\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi,\u003c/sup\u003e n=5. (g) The immunoblotting of UBE2L3, CXCL16 and \u003cem\u003eβ\u003c/em\u003e-actin in lysates, soluble CXCL16 (sCXCL16) in the supernatants (super.) of cultured primary mouse keratinocytes in control group (KC-Ctrl) and in \u003cem\u003eUbe2l3 \u003c/em\u003eknockout group (KC-\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e). (h) Bar plots about the statistics of (g) western blots results. ***，p\u0026lt;0.001, **，p\u0026lt;0.01, ns, not significant, n=3 independent samples. (i) The immunoblotting results of UBE2L3, Pro-IL1β and \u003cem\u003eβ\u003c/em\u003e-actin in lysates, IL-1β p35 and IL-1β p17 in supernatants of KC-Ctrl and KC-\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e. (j) Bar plots about the statistics of (g) western blots results. **，p\u0026lt;0.01, p-value, n=3 independent samples. (k) Mouse control keratinocytes were stimulated with IL-1β(100ng/ml), IL-17A(100ng/ml) for 24 hours. Immunoblotting of STAT3, pSTAT3, CXCL16 and \u003cem\u003eβ\u003c/em\u003e-actin was carried out. (l) The R\u0026amp;D Luminex assay showed that the protein level of CXCL16 was increased in Epi-\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e group. (m) The FPKM of CXCL16 in primary human normal keratinocytes (KC-Ctrl) and primary human normal keratinocytes treated with recombinant human IL-17A cytokines (KC-IL17A) in GSE36287. (n-o) Represent flow cytometric plots and percentage of TNFα and CXCL16 in the epidermis of healthy donors (Epi-HD) and psoriatic patients (Epi-PSO). **，p\u0026lt;0.01, p-value, n=3 independent samples. Epi-HD, n=3, Epi-PSO, n=3. (p) Normal human epidermal keratinocytes (NHKEs) were stimulated with rhIL1β (100ng/ml) and rhIL-17A(100ng/ml) for 24 hours and the STAT3, pSTAT3 and CXCL16 in lysates were measured by western blot. **，p\u0026lt;0.01, *, p\u0026lt;0.05, n=3 independent experiments. The samples were derived from the same experiment, and the gels/blots were processed in parallel.\u003c/p\u003e","description":"","filename":"Picture5.png","url":"https://assets-eu.researchsquare.com/files/rs-5973089/v1/3c083785c8b022264bba7c2b.png"},{"id":77428915,"identity":"e6477de2-8498-4a95-8fba-686a15a08628","added_by":"auto","created_at":"2025-02-28 13:39:55","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":401434,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eCXCL16 was secreted from cDC2 and mDC in human and mouse psoriatic epidermis\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e(a) The single cell profile of myeloid cells about nine classes conserved epidermis across \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e (Mouse-CE) and Ctrl mice (Mouse-UE), delineated by Louvain clustering. MC_Bas, mast cell and basophils. Neu, neutrophils. LC, Langerhans cells. Mac, macrophage. cDC1, type 1 DC. cDC2, type 2 DC. cDC2, type 2 DC. (b) The relative abundance of each myeloid cells types in two groups. (c) The bulk RNA expression of cytokines (Il1b) and chemokine (Cxcl16, Cxcl10 and Adam10) in nine defined classes. Red arrow denotes significance of Cxcl16 in mDC_cDC2 cluster. (d) Pseoudotime analysis of Cxcl16 gene in nine cluster above in myeloid cells. Blue circles indicate the enrichment of Cxcl16 in mDC_cDC2 and Mac. (e) Representative flow cytometric plots and percentage of CXCL16 in mouse DC group in Epi-ctrl and Epi-\u003cem\u003e Ube2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e. Among CXCL16\u003csup\u003e+\u003c/sup\u003eDC, the proportion of CD11b\u003csup\u003e+\u003c/sup\u003eDC (cDC2) and CD103\u003csup\u003e+\u003c/sup\u003eDC (cDC1) in all CXCL16\u003csup\u003e+\u003c/sup\u003eDC. ****，p\u0026lt;0.0001. n=5 independent per group. (f) Schematic strategy for generating \u003cem\u003eUbe2l3 \u003c/em\u003e\u003csup\u003e\u003cem\u003eEpi△\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e-Zbtb \u003c/em\u003e\u003csup\u003e\u003cem\u003e-/-\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e mice. \u003c/em\u003e(g) The phenotype of \u003cem\u003eUbe2l3 \u003c/em\u003e\u003csup\u003e\u003cem\u003eEpi△\u003c/em\u003e\u003c/sup\u003emice and\u003cem\u003e Ube2l3 \u003c/em\u003e\u003csup\u003e\u003cem\u003eEpi△\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e-Zbtb \u003c/em\u003e\u003csup\u003e\u003cem\u003e-/-\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e \u003c/em\u003emice in week 0, 4, and 8. H\u0026amp;E staining of mouse skin in week 8. Epidermal thickness of each group was calculated. **，p\u0026lt;0.01. Immunofluorescence of CD11C(red), K14 (green) and DAPI (blue) , was done in two groups. scale bar= 50 um. (h) Representative flow cytometric plots and the percentage of IL17A\u003csup\u003e+\u003c/sup\u003e in CD3\u003csup\u003e+\u003c/sup\u003eT, γδT and αβT in the epidermis of each group were showed **，p\u0026lt;0.01. ns, not significant. Epi-\u003cem\u003eUbe2l3 \u003c/em\u003e\u003csup\u003e\u003cem\u003eEpi△\u003c/em\u003e\u003c/sup\u003e, n=3, \u003cem\u003eUbe2l3 \u003c/em\u003e\u003csup\u003e\u003cem\u003eEpi△\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e-Zbtb \u003c/em\u003e\u003csup\u003e\u003cem\u003e-/-\u003c/em\u003e\u003c/sup\u003e, n=3. (i) UMAP visualization of different myeloid cells states found in human normal epidermis (NE) and human psoriatic epidermis (PE). pDC, plasm Group composition of\u003c/p\u003e","description":"","filename":"Picture6.png","url":"https://assets-eu.researchsquare.com/files/rs-5973089/v1/10ccf2ebd4f0f1fd109970ae.png"},{"id":77427867,"identity":"c6ecef75-cdd7-41fc-9f0d-7faabae77ba2","added_by":"auto","created_at":"2025-02-28 13:31:55","extension":"png","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":7438971,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eCXCL16 promote Vγ2\u003c/strong\u003e\u003csup\u003e\u003cstrong\u003e+ \u003c/strong\u003e\u003c/sup\u003e\u003cstrong\u003eγδT/ CD8\u003c/strong\u003e\u003csup\u003e\u003cstrong\u003e+\u003c/strong\u003e\u003c/sup\u003e\u003cstrong\u003eT to release IL-17A in psoriatic epidermis and inhibition of CXCL16 alleviated psoriasis-like lesion\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e(a) The percentage of Vγ2\u003csup\u003e+\u003c/sup\u003e γδT in γδT in Der-Ctrl and Der-\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e group. *, p\u0026lt;0.05. (b) Schematic strategy for studying the migration of Vγ2\u003csup\u003e+\u003c/sup\u003e γδT cells with or without Cxcl16 for 24 hours. (c) The migrated cell was viewed by immunofluorence staining with DAPI (blue), five independent views of lower chambers of insects were calculated between Der-Ctrl and Der-\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e group. *, p\u0026lt;0.05. (d) The pseudotime analysis in T lymphocytes in eight groups. CD8T, DETC, NK, ILCs, Unknown, Treg, gdT17, Cyc-T and CD4T. The green circle indicates DETC and the purple circle indicates gdT17. (e) Schematic strategy for studying CD3\u003csup\u003e+\u003c/sup\u003eT cells, sorted by flow cytometry and stimulated with or without rmCXCL16 (100ng/ml). Then flow cytometry was carried out to study the percentage of KI67 and IL-17A in CD3\u003csup\u003e+\u003c/sup\u003eT cells. (h) \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi \u003c/sup\u003emice were treated with isotype antibodies or anti-CXCL16 mAb. H\u0026amp;E staining of back skin and ear skin was performed. **, p\u0026lt;0.01. (j) Epidermal single cells were stimulated with or without recombinant human CXCL16 and flow cytometric analysis of IL-17A was showed. (k) The percentage of IL-17A\u003csup\u003e+ \u003c/sup\u003eCD3\u003csup\u003e+\u003c/sup\u003eT /CD3\u003csup\u003e+\u003c/sup\u003eTcells, CD8\u003csup\u003e+\u003c/sup\u003eIL-17A\u003csup\u003e+\u003c/sup\u003e/IL-17A\u003csup\u003e+\u003c/sup\u003e T cells, CD4\u003csup\u003e+\u003c/sup\u003eIL-17A\u003csup\u003e+\u003c/sup\u003e/IL-17A\u003csup\u003e+\u003c/sup\u003eT cells in isotype cytokine and rhCXCL16 cytokine. *, p\u0026lt;0.05, **, p\u0026lt;0.01. (l) Flow cytometric analysis of the percentage of Ki-67\u003csup\u003e+\u003c/sup\u003e IL-17A\u003csup\u003e+ \u003c/sup\u003eCD8\u003csup\u003e+\u003c/sup\u003e in CD8\u003csup\u003e+ \u003c/sup\u003eJurkat cells. (j) and proliferation activity of CD8\u003csup\u003e+ \u003c/sup\u003eJurkat cells.\u003c/p\u003e","description":"","filename":"Picture7.png","url":"https://assets-eu.researchsquare.com/files/rs-5973089/v1/2e282245e5fac285657c1604.png"},{"id":77427856,"identity":"556ee334-1440-4f94-a75c-a01892b52877","added_by":"auto","created_at":"2025-02-28 13:31:55","extension":"png","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":1125815,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eGraphical abstract\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eA psoriasis-like lesion mouse model was generated by conditional knock out \u003cem\u003eUbe2l3\u003c/em\u003e in epidermis (\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e), which was compared with human psoriasis scRNA-seq data and facilitate cross-species comparisons of differentiation dynamics and ligand-receptor pathways in epidermis. In particular, IL-17A was regulated by CXCR6\u003csup\u003e+\u003c/sup\u003e Vγ2\u003csup\u003e+ \u003c/sup\u003eγδT in mouse while CXCR6\u003csup\u003e+ \u003c/sup\u003eCD8\u003csup\u003e+ \u003c/sup\u003eT in human. \u003cem\u003eUbe2l3\u003c/em\u003e reduction in keratinocytes activated IL-1β and then promote CXCL16 expression through STAT3 signaling. CXCR6\u003csup\u003e+\u003c/sup\u003eγδT17/ Tc17cells are prevalent within the epidermal immune microenvironment of psoriasis. Deletion of CXCL16 inhibits the progression of psoriasis-like lesions in \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e \u003c/em\u003emice. (Created by biorender.com)\u003c/p\u003e","description":"","filename":"Picture8.png","url":"https://assets-eu.researchsquare.com/files/rs-5973089/v1/4fa504bf1ddd5f6b2d18febc.png"},{"id":93464593,"identity":"1509255f-405e-439e-a1be-2d6b4723918c","added_by":"auto","created_at":"2025-10-14 07:08:23","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":31233697,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-5973089/v1/db5c6032-ba45-420e-8eca-e21a5aa46a39.pdf"},{"id":77427859,"identity":"f84f7664-710b-4cb7-85f3-d4ca602bae3a","added_by":"auto","created_at":"2025-02-28 13:31:55","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":6151678,"visible":true,"origin":"","legend":"Supplementary information","description":"","filename":"Supplementaryinformation.pdf","url":"https://assets-eu.researchsquare.com/files/rs-5973089/v1/4200abb55b76f7e7576a3094.pdf"}],"financialInterests":"There is \u003cb\u003eNO\u003c/b\u003e Competing Interest.","formattedTitle":"Cell atlas of human psoriasis and epidermal specific Ube2l3 deficiency mouse highlighting CXCL16/CXCR6 orchestrating the development of psoriasis","fulltext":[{"header":"INTRODUCTION ","content":"\u003cp\u003ePsoriasis is a chronic, complex immune-mediated inflammatory disorder with cutaneous and systemic manifestations\u003csup\u003e\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u003c/sup\u003e. Its pathogenesis is characterized by the involvement of keratinocytes, dendritic cells and T cells, which play a central role in the disease process\u003csup\u003e\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e\u003c/sup\u003e. To facilitate research on the mechanisms of action that drive inflammatory and autoimmune processes associated with psoriasis, it would be highly beneficial to have a suitable animal model\u003csup\u003e\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e, \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e\u003c/sup\u003e. Moreover, keratinocytes may be a crucial factor in both the onset and perpetuation of psoriasis\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e. Our and other previous research have indicated that diminished Ubiquitin Conjugating Enzyme E2 L3 (UBE2L3) expression is linked to elevated Interleukin 1β (IL-1β) production in the epidermis of psoriasis.\u003csup\u003e\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e, \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e\u003c/sup\u003e. The epidermal immune environment is of pivotal importance in the pathogenesis of psoriasis\u003csup\u003e\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e\u003c/sup\u003e. Furthermore, UBE2L3 interacts with the ubiquitin enzyme 3, including TRIP12 and AREL1, to facilitate the turnover of the precursor IL-1β (pro-IL-1β) \u003csup\u003e\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e\u003c/sup\u003e. This process is of critical importance to the functioning of the immune system. Consequently, we firstly established a mouse model of epidermal \u003cem\u003eUbe2l3\u003c/em\u003e deficiency in order to gain further insight into the pathogenesis of psoriasis. This mouse model offers a valuable opportunity to explore the pathogenesis of psoriasis\u003csup\u003e\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e, \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u003c/sup\u003e. It is of great importance to compare the mutant mice to psoriatic patient samples at the level of bulk RNA, single cell RNA, and protein.\u003c/p\u003e \u003cp\u003eIn contrast to adaptive IL-17-producing T cells, dermal IL-17-producing γδT (γδT17) cells are programmed for the earliest IL-17 response to various pathogens\u003csup\u003e\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e\u003c/sup\u003e. In these settings, γδT17 cells have been demonstrated to respond to cytokines, predominantly IL-23 and IL-1β\u003csup\u003e12\u003c/sup\u003e. Vγ2 T (IL-17A producers,Heilig \u0026amp; Tonegawa nomenclature) cells have been demonstrated to contribute to the pathogenesis of autoimmune diseases by through the production of IL-17A\u003csup\u003e\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e\u003c/sup\u003e. Integration of single cell sequencing (scRNA-seq) data and spatial-seq transcriptomic CD8\u003csup\u003e+\u003c/sup\u003eIL17\u003csup\u003e+\u003c/sup\u003eT (Tc17) enabled the definition of Tc17, which was identified as a principal source of IL-17A in psoriatic skin\u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e, \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e\u003c/sup\u003e. However, the function of γδT17 and Tc17 in epidermal psoriatic lesions remains to be elucidated.\u003c/p\u003e \u003cp\u003eChemokine (C-X-C motif) ligand 16 (CXCL16) functions as a ligand for C-X-C chemokine receptor type 6 (CXCR6), and both are all upregulated in psoriasis. They mediate cutaneous recruitment of human CD8\u003csup\u003e+\u003c/sup\u003eT cells\u003csup\u003e\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e, \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e\u003c/sup\u003e. The expression of CXCR6 has been observed in Th17 cells and Tc17 with increased levels of CXCL16\u003csup\u003e18\u003c/sup\u003e. Furthermore, CXCL16 has been demonstrated to regulate the migration of CXCR6-expressing CD8\u003csup\u003e+\u003c/sup\u003e T cells and promote Tc17 differentiation\u003csup\u003e\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e, \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e\u003c/sup\u003e. Additionally, CXCL16 is secreted by macrophages and dendritic cells (DCs) to attract memory T cells\u003csup\u003e\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e\u003c/sup\u003e. Molecular studies demonstrated that fibrosus cells exposed to IL-1\u0026szlig;, exhibited a notable increase in CXCL16 expression compared to control cells\u003csup\u003e\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e\u003c/sup\u003e. The precise function of CXCL16 in dendritic cells (DCs) and CXCR6 in γδT and CD8\u003csup\u003e+\u003c/sup\u003e T cells, particularly within the context of the epidermal psoriatic milieu, remains to be elucidated.\u003c/p\u003e \u003cp\u003eIn this study, we demonstrated an epidermal \u003cem\u003eUbe2l3\u003c/em\u003e conditional knockout mouse model exhibited a complete mimicry of the human psoriatic phenotype. This was achieved through the analysis of using bulk RNA levels, protein levels and single-cell RNA sequencing (scRNA-seq). In conclusion, UBE2L3 functions as a keratinocyte-intrinsic suppressor of epidermal IL-17 production in mouse Vγ2\u003csup\u003e+\u003c/sup\u003e γδT and human CD8\u003csup\u003e+\u003c/sup\u003e T cells via the CXCL16/CXCR6 signaling pathway in psoriasis.\u003c/p\u003e"},{"header":"RESULTS","content":"\u003cp\u003e \u003cb\u003eCutaneous phenotypes of epidermal deficiency of Ube2l3 mouse models exhibit a high degree of similarity to that observed in human psoriasis\u003c/b\u003e \u003c/p\u003e \u003cp\u003ePreviously, our research demonstrated that in patients with psoriasis, UBE2L3 is predominantly downregulated in the epidermis, with the exception of the dermis\u003csup\u003e\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e\u003c/sup\u003e. To gain further insight into the underlying mechanisms, we established a Ube2l3 epidermal deficiency mouse model (\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e, Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ea). The cutaneous phenotype of \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e murine models exhibited notable erythema, thickness, and scales as the induction time by tamoxifen (TAM) was prolonged (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eb-d). The substantial downregulation of \u003cem\u003eUbe2l3\u003c/em\u003e in the epidermis of \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e group was substantiated through the implementation of western blot, bulk RNA sequencing, immunofluorescence and immunohistochemistry staining (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ee, \u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ef, Fig.\u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003ea). A typical psoriasis-like skin phenotype was established in back, ear and tail (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eg), measured by significantly increased epidermal thickness in \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e mice (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eh, \u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ei, and \u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ej), comparable to human psoriatic skin lesions (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ek). The increased angiogenesis (CD31), proliferation (Ki67) and immunocytes infiltration (CD3, CD11c), which corresponded to the psoriasis-like phenotype and inflammatory stage (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003el, Fig.\u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003eb-d), thus provide a new insight into the ability to mimic psoriasis disease in the mouse model.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003eA comparative analysis of RNA and protein levels between\u003c/b\u003e \u003cb\u003eUbe2l3\u003c/b\u003e\u003csup\u003e△Epi\u003c/sup\u003e \u003cb\u003emice and human psoriasis skin\u003c/b\u003e\u003c/p\u003e \u003cp\u003eTo investigate the molecular mechanisms involved in sustaining skin inflammation in the epidermis or dermis, we conducted ex vivo bulk-RNA sequencing and Illumina array analysis of epidermis and dermis respectively from \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e mice and control (Ctrl) mice. The principal component analysis (PCA) demonstrated a distinct separation of the four groups (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ea). The number of genes in the dermis is significantly lower than in the epidermis when using the same p-value threshold of less than 0.05 and an absolute log2 fold change greater than 1.5 are employed (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eb, \u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ec, and \u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ed). The data were obtained from GSE166388, GSE68937 and GSE68923, and a comparison was made between human psoriatic lesional skin and normal skin from controls in the epidermis (Fig. S2a-b). The genes that were found to be upregulated were collated and analysed by using the Metascape website\u003csup\u003e\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e\u003c/sup\u003e in order to identify all molecules and enrichment pathways. The \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e mouse was found to be closely associated with human psoriasis in the epidermis, specifically in relation to the immune response (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ee-f). The enrichment analyses of the dermis, conducted using differentially expressed genes (DEGs), demonstrated a lesser degree of correlation with psoriasis compared to the epidermis (Fig.S2c-d). The genes found to be related to psoriasis belong to the majority of members of the cytokine and chemokine families, including interleukin-1 (IL-1), IL-17, IL-6/IL-23\u003csup\u003e25\u003c/sup\u003e, chemokines and their receptors, antimicrobial peptides (AMPs), tumour necrosis factor (TNF), toll-like receptors (TLRs), as well as representative Type I interferon receptors (IFNRs). The heatmap indicated that these families were all significantly upregulated in the epidermis of \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e mice, which is analogous to human psoriasis (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eg, \u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eh). The Illumina array analysis revealed increase in protein levels of AMPs (S100A8, S100A9), vascular endothelial growth factor (VEGF), chemokines (CCL2, CCL22, CXCL2), and cytokine (IL-17A) in the epidermis of the \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e mouse (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ei). Furthermore, the iTRAQ analysis revealed an increase in antimicrobial peptides (AMPs), while the Illumina array analysis indicated a tendency towards an increase in tumour necrosis factor (TNF), interleukin 12/interleukin 23p40 (IL-12/IL23 P40), and IL-17A in the human dermis (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ej). However, no significant differences were observed in psoriasis-related families in the dermis as that in the epidermis, as indicated by both the enrichment analysis and the heatmap (Fig. S2c-d). Consequently, a comparison of RNA and protein levels between the \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e mice and human psoriasis skin revealed a striking similarity.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eMouse versus human epidermal lymphocytes subpopulations with respect to γδT and CD8\u003csup\u003e+\u003c/sup\u003eT cells secreting IL-17A\u003c/h2\u003e \u003cp\u003eWe next sought to compare subpopulations within the T lymphocytes compartments from human psoriasis skin lesion and \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e mouse model. IL-17A, the principal downstream cytokine of IL-23, is most strongly implicated in the pathogenesis of psoriasis, with T lymphocytes secreting it in the greatest quantity\u003csup\u003e\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e\u003c/sup\u003e. scRNA-seq data showed a prevalence of infiltrating IL-17A secreting γδT (gdT17) cells in the epidermis of \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e mice (UE), whereas dendritic epidermal t cells (DETC) were dominant in the epidermis of control mice (CE) (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ea, \u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eb). The gdT17 emonstrated a high level of expression of \u003cem\u003eIl17a, Il17f\u003c/em\u003e and \u003cem\u003eTcrg-V4\u003c/em\u003e, whereas the DETC displayed a high relative expression of \u003cem\u003eCd3e, Trdv4, Tcrg-v5\u003c/em\u003e and \u003cem\u003eFcer1g\u003c/em\u003e\u003csup\u003e\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e\u003c/sup\u003e. Both cell types displayed a lack of \u003cem\u003eCd4/Cd8a\u003c/em\u003e expression (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ec). Furthermore, Immune CellAI-mouse\u003csup\u003e\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e\u003c/sup\u003e was employed to elucidate the infiltration score in various cell types in both the epidermis and dermis. The findings revealed that the infiltration of γδT was markedly more pronounced in the epidermis of \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e mice in comparison to the dermis (Fig.S3a).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eIn the historical context, IL-17-producing T cells have been predominantly regarded as CD4\u003csup\u003e+\u003c/sup\u003e T cells. However, recent observations have provided growing evidence for the presence of IL-17 producing CD8\u003csup\u003e+\u003c/sup\u003e T cells in psoriasis\u003csup\u003e\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e\u003c/sup\u003e. A similar isolation of human lymphocytes yielded 15 clusters, including IL-17 signalling-related CD8\u003csup\u003e+\u003c/sup\u003e T cells (CCL20_Tc17, IL-17A_Tc17, IL-17F_Tc17 and IL26_Tc17), based on expression of \u003cem\u003eCD4\u003c/em\u003e, \u003cem\u003eCD8A\u003c/em\u003e, or \u003cem\u003eCD8B (\u003c/em\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ed,\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ee,\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ef\u003cem\u003e)\u003c/em\u003e, as observed in other recent studies\u003csup\u003e\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e\u003c/sup\u003e. Moreover, Th17/Tc17 cytokines, including \u003cem\u003eIL17A\u003c/em\u003e, \u003cem\u003eIL17F\u003c/em\u003e, \u003cem\u003eIL26\u003c/em\u003e\u003csup\u003e\u003cem\u003e31\u003c/em\u003e\u003c/sup\u003e,CCL20 and \u003cem\u003eCXCL13\u003c/em\u003e\u003csup\u003e32\u003c/sup\u003e were also employed (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ed-\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ef, and Fig.S3b). Another group was defined as cytotoxic T cells (Tc), which were identified by the presence of \u003cem\u003eGNLY\u003c/em\u003e, \u003cem\u003eGZMH\u003c/em\u003e, \u003cem\u003eHSPA1B\u003c/em\u003e and \u003cem\u003eIFNG\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ed-f). The simultaneous up-regulation of the T cell receptor signalling pathway, Th17 cell differentiation and Th1 and Th2 cell differentiation was observed in both the mouse gdT17 and human Tc17 clusters (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eg, h). The nuclear receptor retinoid-related orphan receptor gamma t (\u003cem\u003eRORγt, RORC\u003c/em\u003e) has been identified as a pivotal regulator of the differentiation and expansion of Th17 cells\u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e. One of the most significant findings was that Rorc, a transcription factor (TF) with a pivotal role in the differentiation and expansion of Th17 cells, was identified as one of the top TFs in the mouse gdT17 cluster (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ei). This observation aligns with the characteristics of human psoriasis type 17 cell differentiation\u003csup\u003e\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e\u003c/sup\u003e. We proceeded to perform flow cytometry on IL-17A source cells derived from \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e mice skin lesions and psoriasis patients skin biopsies, in comparison with those from site-matched control mice (Ctrl)/ \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e mice. An increase in IL-17A-producing γδT cells was observed in the epidermal samples of \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e mice (Epi-\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e mice samples) in comparison with those of control mice (Epi-Ctrl). This was in contrast to the observed increase in TNFα secretion (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ej, Fig.S3c). In addition, an increase in IL-17A-producing CD8\u003csup\u003e+\u003c/sup\u003e T cells (Tc17) was observed in epidermal psoriatic lesions (PSO-Epi) (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ek).\u003c/p\u003e \u003cp\u003eFollowing dermal conversion, no notable discrepancy in IL-17A secretion was discerned between the \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e mice samples (Der-\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e) and the control mice (Der-Ctrl) (Fig.S3d). The administration of an antibody that neutralises IL-17A to \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e mice during the effector phase resulted in a reduction in epidermal thickness, a decrease in the infiltration of lymphocytes (CD3), and an enhanced tendency for epidermal keratinocytes to differentiate (Loricrin) (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003el,\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003em and \u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003en). Therefore, the \u003cem\u003eUbe2l3\u003c/em\u003e conditional deficiency model of psoriasis in epidermal γδT cells is largely analogous to the IL-17A-secreting CD8\u003csup\u003e+\u003c/sup\u003eT cells observed in human psoriasis.\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eCrosstalk with CXCL16-CXCR6 signaling informed ligand receptor analysis\u003c/h3\u003e\n\u003cp\u003eThe maintenance of homeostasis requires intercellular communication between diverse cell types. An approach to assess such communication is provided by single-cell integration. The initial step involved the tabulation of inferred ligand-receptor interactions across keratinocytes, myeloid cells (myeloids) and NK/T cells (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ea, \u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eb, \u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ec). Subsequently, the most highly expressed ligand-receptor pairs from known major pathways operating in mouse epidermal skin were curated. Within the epidermis, one of the most prominent pairs of NK/T cells demonstrated crosstalk with myeloid cells or keratinocytes through Cxcl16-Cxcr6 signalling (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ed-e). Cxcr6 was previously demonstrated to be expressed in T cells, which were predominantly enriched in gdT17 clusters with high expression of Vγ2 and Vγ4 specified markers simultaneously (\u003cem\u003eTcrg-V4, 5830411N06Rik and Cd163l\u003c/em\u003e)\u003csup\u003e\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e, \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e\u003c/sup\u003e(Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ef). A greater proportion of IL-17A and CXCR6 was observed to be co-expressed in Epi-\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e, as evidenced by flow cytometry analysis (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eg). The percentage of Vγ2\u003csup\u003e+\u003c/sup\u003eγδT cells within the γδT cell population was found to be significantly increased, while the frequency of Vγ3\u003csup\u003e+\u003c/sup\u003e cells was observed to be abnormally decreased (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eh). The observed up-regulation of IL-17A\u003csup\u003e+\u003c/sup\u003e Vγ2\u003csup\u003e+\u003c/sup\u003eγδT cells indicates that the increase in IL-17A proportion in T cells is driven by Vγ2\u003csup\u003e+\u003c/sup\u003eγδT upregulation (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ei). The intraperitoneal neutralisation of Vγ2 in \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e psoriasis-like mice has led to the conclusion that Vγ2\u003csup\u003e+\u003c/sup\u003eγδT may play an important role in this psoriasis-like mouse model. Furthermore, a decreased trend towards psoriasis-like lesions, epidermal thickness (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ej-k) and IL-17A proportion of γδT in the epidermis of the anti-Vγ2 group (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003el, Fig. S4a) was observed. No significant difference was observed between the dermal IL-17A and TNFα proportions in the isotype and anti-Vγ2 groups (Fig.S4b). In addition, the knock-out of \u003cem\u003eRag1\u003c/em\u003e in \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e mice (\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e-Rag1\u003c/em\u003e \u003csup\u003e\u003cem\u003e\u0026minus;/\u0026minus;\u003c/em\u003e\u003c/sup\u003e) also exhibited a decreased epidermal thickness compared \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e mice (Fig. S4c). These all indicate that Vγ2\u003csup\u003e+\u003c/sup\u003eγδT-secreting IL-17A played a pivotal role in the inflammatory environment of the mouse epidermis.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eWe next ascertain the relevance of human psoriasis in epidermal CXCR6\u003csup\u003e+\u003c/sup\u003eT cells. The expression of CXCR6 found to be elevated in psoriatic lesional skin at the baseline stage (LS-baseline) in comparison to non-lesional skin (NL) and following the administration of biologic treatment (LS-treated) (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003em). A comparison was made between lymphocytes isolated from both adult epidermal healthy skin and epidermal psoriasis samples from the scRNA-seq dataset. The spatial transcriptomics, dotplot analysis and heatmap in the scRNA-seq analysis revealed an abundance of CXCR6 enrichment in the Tc17 population in the epidermis (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003en-o, Fig.S4d-e), which is consistent with the immunofluorescence staining and flow cytometry analysis in the epidermis in human psoriasis (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ep-q). In conclusion, epidermal IL-17A signalling was demonstrated to be regulated by CXCR6\u003csup\u003e+\u003c/sup\u003e Vγ2\u003csup\u003e+\u003c/sup\u003e γδ T cells in the mouse model and by CXCR6\u003csup\u003e+\u003c/sup\u003e CD8\u003csup\u003e+\u003c/sup\u003e T cells in the human subject.\u003c/p\u003e\n\u003ch3\u003eIL-17A and IL-1β stimulated secretion of CXCL16 in keratinocytes through STAT3 signaling\u003c/h3\u003e\n\u003cp\u003eThe scRNA-seq analyses of the \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e psoriasis-like mouse model provided evidence to support the hypothesis that keratinocyte subpopulation dysfunction may play a role in the pathogenesis of psoriasis. The proportion of basal and spinous cells between healthy and psoriatic skin was analysed, revealing an increase in both proliferating basal cells (prolif.Basal), specified by \u003cem\u003eTop2a, Mki67\u003c/em\u003e\u003csup\u003e36\u003c/sup\u003e, and cycling spinous cells (Spinous II) in the \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e mouse epidermis (Mouse-UE) compared to the control epidermis (Mouse-CE). This suggests that the basal layer exhibited hyperproliferation (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ea-b). We therefore conducted a detailed investigation into the up-regulated expression of CXCL16 in Mouse-UE, which was predominantly expressed in the prolif.Basal cluster (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ed-e). This finding was consistent with the CXCL16 expression observed in CD45- cells, as analysed by flow cytometry (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ef). We therefore proceeded to investigate the relationship between Ube2l3 deficiency and CXCL16 secretion.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003ePrevious studies have demonstrated that the overexpression of UBE2L3 reduces its binding to TRIM21, consequently leading to a decrease in STAT3 pathway activity and a reduction in the level of the IL-1β precursor (pro-IL-1β). Furthermore, molecular studies demonstrated that annulus fibrosus cells exposed to IL-1β, but not TNF-α, exhibited a notable elevation in CXCL16 expression relative to control cells\u003csup\u003e\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e\u003c/sup\u003e. The results demonstrated that both CXCL16 and IL-1β were elevated in \u003cem\u003eUbe2l3\u003c/em\u003e-deficient keratinocytes (KC-\u003cem\u003eUbe2l3\u003c/em\u003e△) relative to control cells (KC-Ctrl) (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eg-i). Upon stimulation of keratinocytes with or without recombinant mouse IL-17A (rmIL-17A) and recombinant IL-1β (rmIL-1β), the protein level of CXCL16 was markedly increased, exhibiting a similar trend to that of STAT3 signalling (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ek). Furthermore, CXCL16 was found to be upregulated in epidermal \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e mice (\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u0026minus;\u003c/sup\u003eEpi) (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003el). In human keratinocytes stimulated with IL-17A, an increase in CXCL16 mRNA levels was observed (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003em). Similarly, the same up-regulated tendency of CXCL16 in psoriasis in epidermis (PSO-Epi) was observed in CD45- cells by flow cytometry (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003en-o). Upon stimulation of normal human epidermal keratinocytes (NHEKs) with IL-1β and IL-17A, a significant up-regulation of CXCL16 was observed in lysates, accompanied by activation of the STAT3 pathway (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ep). In conclusion, the data collectively indicate that CXCL16 is secreted by \u003cem\u003eUbe2l3\u003c/em\u003e-deficient keratinocytes and stimulated by IL-1β and IL-17A. This suggests that CXCL16 may act as a chemoattractant for T cells, which in turn secrete IL-17A, forming a positive feedback loop in psoriatic keratinocytes.\u003c/p\u003e\n\u003ch3\u003eUp-regulated CXCL16 secreted by cDC2/mDC controlling CXCR6 γδT/CD8T IL-17A secretion in psoriatic epidermis\u003c/h3\u003e\n\u003cp\u003escRNA-seq and bulk RNA sequencing analysis indicated the potential involvement of CXCL16-CXCR6 signalling between myeloid cells and natural killer (NK)/T cells in the epidermis of \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e psoriasis-like mice, with no evident impact on the dermis (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ed-e, Fig.S5a-c). Therefore, we proceeded to evaluate the proportion of CXCL16 in myeloid cells, identifying a notable presence in the migrating dendritic cell (mDC) and cDC2 (type 2 DC) population, specifically marked by \u003cem\u003eItgam, Irf4, Fscn1, Cacnb3 and Ccr7\u003c/em\u003e\u003csup\u003e37\u003c/sup\u003e(Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ea-c, Fig.S5e). The results of the pseudo-time analysis indicated that Cxcl16 was predominantly expressed in mDC_cDC2 and Mac (macrophage) (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ed). The same trend was corroborated by flow cytometry, which demonstrated that cDC2 was the predominant cell type secreting CXCL16 in the epidermis (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ee). Following the knockout of the Zbtb gene in \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e mice, \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e-Zbtb\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u0026minus;/\u0026minus;\u003c/em\u003e\u003c/sup\u003e mice were generated after TAM induction (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ee). The epidermal thickness of the psoriasis-like lesions was found to be decreased in conjunction with the down-regulation of CD11c and keratin 14 (K14) (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ef-g). Furthermore, we observed a notable reduction in the proportion of γδT cells secreting IL-17A in \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e-Zbtb\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u0026minus;/\u0026minus;\u003c/em\u003e\u003c/sup\u003e(Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eh). In a human myeloid single-cell analysis, a comparable population of mDC and cDC2 was identified based on the expression of \u003cem\u003eCCR7, LAMP3, CLEC10A, CD1C\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e, \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e\u003c/em\u003e\u003c/sup\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ej, Fig.S5f-g), The results demonstrated that CXCL16 was markedly elevated in the epidermis of PSO (Epi-PSO) specimens, as evidenced by heatmap analysis and flow cytometry (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ei-k). Similarly, a comparable trend is observed in myeloid cells, with the involvement of analogous pathways, such as the regulation of cell activation (Fig.S5h-i). Therefore, CXCL16 was released by the cDC2/mDC population simultaneously and may be involved in the regulation of γδT17 secretion in the psoriatic epidermis.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e\n\u003ch3\u003eCXCL16 facilitates Vγ2 γδT/ CD8T migration, proliferation and IL-17A release in psoriatic epidermis\u003c/h3\u003e\n\u003cp\u003eIn wild-type mice, Vγ2\u003csup\u003e+\u003c/sup\u003e γδT was observed to be absent in the epidermis and present in the dermis\u003csup\u003e\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e\u003c/sup\u003e. In psoriasis-like \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e mice, the percentage of Vγ2\u003csup\u003e+\u003c/sup\u003e γδT cells was significantly increased in the dermis (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003ea). Epithelial cells lining the mucosal surfaces play a crucial role in this process by producing specific chemokines, such as CXCL16, which serve as attractants for γδT cells expressing the corresponding chemokine receptors\u003csup\u003e\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e\u003c/sup\u003e. Furthermore, IL-23R signalling is responsible for the generation of CXCR6\u003csup\u003e+\u003c/sup\u003e cells from IL-17\u003csup\u003e+\u003c/sup\u003e SLAMF6\u003csup\u003e+\u003c/sup\u003e cells\u003csup\u003e42\u003c/sup\u003e. It was therefore postulated that there is a relationship between CXCL16 and CXCR6\u003csup\u003e+\u003c/sup\u003eTc17/γδT17. The migration assay demonstrated that CXCL16 exerted a migratory effect on dermal Vγ2\u003csup\u003e+\u003c/sup\u003e γδT (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003eb-c). In accordance with the limited research on the conversion of DETC to Vγ2\u003csup\u003e+\u003c/sup\u003e γδT17, we conducted a pseudotime analysis that revealed the non-near differentiation of Vγ2\u003csup\u003e+\u003c/sup\u003e γδT17 and DETC (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003ed). Next, epidermal and dermal Vγ2\u003csup\u003e+\u003c/sup\u003e γδT cells were sorted and stimulated with Cxcl16 (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003ee). The results demonstrated that the percentage of Ki-67 and IL-17A was increased in two-week-old TAM-induced \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e mice (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003ef). Furthermore, the secretion of IL-17A was increased in Vγ2\u003csup\u003e+\u003c/sup\u003e γδT cells in 6 week-TAM induced \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e mice (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003eg). CXCL16 neutralization in \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e mice resulted in a reduction in the epidermal thickness of psoriasis-like lesions (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003ei). Upon stimulation of the human epidermal psoriasis epidermis (Human-Epi-CD3\u003csup\u003e+\u003c/sup\u003eT) with recombinant human CXCL16 (rhCXCL16), an increase in the percentage of IL-17A secretion was observed in CD3\u003csup\u003e+\u003c/sup\u003eT cells, particularly in CD8\u003csup\u003e+\u003c/sup\u003eT cells (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003ej-k). In wild-type (WT) mice, it was observed that recombinant mouse CXCL16 (rmCXCL16) injection resulted in an increase in ear epidermal thickness, in comparison to rmIL-23 injection in the right ear (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003el, \u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003em and \u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003en). Therefore, CXCL16 can facilitate the migration, proliferation and IL-17A secretion of Vγ2\u003csup\u003e+\u003c/sup\u003e γδT cells in mouse epidermis and CD8\u003csup\u003e+\u003c/sup\u003eT cells in human psoriatic epidermis.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e"},{"header":"DISCUSSION","content":"\u003cp\u003eIn this study, scRNA-seq from human psoriasis skin was integrated to map subpopulation composition and intercellular communication within the major compartments of the skin (keratinocytes, myeloid cells and NK/T cells), thereby generating a comprehensive resource for hypotheses regarding the epidermal immune environment in psoriasis. A psoriasis-like lesion mouse model was generated, which may facilitate further insights into the pathogenesis of psoriasis\u003csup\u003e\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e\u003c/sup\u003e. The data were compared with mouse skin scRNA-seq data to facilitate cross-species comparisons of differentiation dynamics and ligand-receptor pathways. In particular, the CXCL16-CXCR6 signalling pathway was identified as a highly active cluster in keratinocytes/myeloids and NK/T cells (Graphical abstract).\u003c/p\u003e \u003cp\u003eSome research has indicated that pathogenic CXCR6\u003csup\u003e+\u003c/sup\u003e Th17 populations are induced in autoimmunity\u003csup\u003e\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e\u003c/sup\u003e. In the context of inflammatory diseases, the activation of CXCR6\u003csup\u003e+\u003c/sup\u003e T cells has been identified as a key factor in the differentiation of T cells into Th17 and the subsequent secretion of pro-inflammatory cytokines\u003csup\u003e\u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e\u003c/sup\u003e. In human, CXCR6-expressing cells were absent in the epidermis of healthy skin\u003csup\u003e\u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e\u003c/sup\u003e. In contrast, CXCR6\u003csup\u003e+\u003c/sup\u003e CD8\u003csup\u003e+\u003c/sup\u003e T cells were significantly increased in both peripheral blood and skin in psoriasis patients\u003csup\u003e\u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e\u003c/sup\u003e. In mouse model, previous sltudies have demonstrated that dermal γδT cells are the primary IL-17-producing cells in the skin that respond to IL-23 stimulation in an IMQ (imiquimod) induced psoriasis-like mouse model\u003csup\u003e\u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e45\u003c/span\u003e\u003c/sup\u003e. However, there is a paucity of research examining the epidermal CXCL16-CXCR6/IL-17 signalling pathway in psoriasis patients and psoriasis-like mouse models.\u003c/p\u003e \u003cp\u003eIn addition to it, IL-17-induced effector cells in skin inflammation are keratinocytes\u003csup\u003e\u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e\u003c/sup\u003e. In our studies, we found that CXCL16 was secreted by \u003cem\u003eUbe2l3\u003c/em\u003e -deficient keratinocytes, stimulated by IL-1β through the STAT3 pathway, together with CXCL16 from cDC2 and mDC, promoted the migration and proliferation of γδT17 cells in the epidermis of \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e mice. The same activated pathway was found in human psoriasis epidermis and the key cell was CD8\u003csup\u003e+\u003c/sup\u003e Tc17 (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). We first compared the psoriasis-like mouse model and human psoriasis patients in the epidermis and found both CXCL16-CXCR6 signaling played a crucial role in psoriatic cell communication. Our single cell trajectory analysis shed the light on the root of convert and demonstrated the reduced potential of DETC to Vγ2\u003csup\u003e+\u003c/sup\u003e γδT17 (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003ed). The increased epidermal Vγ2\u003csup\u003e+\u003c/sup\u003e γδT17 was derived from the dermis and CXCL16 from keratinocytes/dendritic cells may further facilitate Vγ2\u003csup\u003e+\u003c/sup\u003e γδT17 proliferation and differentiation (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003ef-g). We also found that cutaneous injection of CXCL16 neutralising antibody contributed to the amelioration of psoriasis-like lesion in \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e mice (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003ei). Therefore, CXCL16 may be another therapeutic target and be suitable for use in a precision medicine approach to psoriasis treatment, especially in patients with elevated CXCL16.\u003c/p\u003e \u003cp\u003eOverall, our findings indicate that CXCR6\u003csup\u003e+\u003c/sup\u003eγδT17/ Tc17cells are prevalent within the epidermal immune microenvironment of psoriasis \u003csup\u003e\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e, \u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e\u003c/sup\u003e. The elevated level of CXCL16 produced by \u003cem\u003eUbe2l3\u003c/em\u003e-deficient serves to augment the differentiation and recruitment of γδT17/ Tc17cells, thereby establishing a positive feedback loop that facilitates keratinocyte advancement in psoriasis. Furthermore, we demonstrated that the deletion of CXCL16 inhibits the progression of psoriasis-like lesions in \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e mice and exhibits a therapeutic effect in vivo. The findings of our study offer valuable insights that may inform the development of a therapeutic approach utilising anti-CXCL16 neutralising antibodies, with the potential to provide alleviation of psoriasis.\u003c/p\u003e"},{"header":"METHODS","content":"\u003cdiv id=\"Sec10\" class=\"Section2\"\u003e \u003ch2\u003eMice and ethics\u003c/h2\u003e \u003cp\u003eAll participating patients with plaque psoriasis patients and healthy donors gave written informed consent before inclusion. A skin biopsy with a diameter of 1\u0026ndash;2 cm was taken from healthy donors\u0026rsquo; or psoriasis patients\u0026rsquo; extremities and/or back. The study was approved by the Ethics Committee of the Second Affiliated Hospital, Zhejiang University School of Medicine (2020-No.135).\u003c/p\u003e \u003cp\u003eIn the animal study, C57BL/6 mice was supplied by SLAC Laboratory Animal Co. (Shanghai, China), \u003cem\u003eZbtb46-DTR\u003c/em\u003e mice (Jackson Laboratories), \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003efl/fl\u003c/em\u003e\u003c/sup\u003e (Cyagen)and were housed under specific pathogen-free (SPF) conditions. \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003efl/fl\u003c/em\u003e\u003c/sup\u003e (mice were crossed to K14-Cre\u003csup\u003eERT\u003c/sup\u003e mice (Jackson Laboratories) to generate \u003cem\u003eUbe2l3\u003c/em\u003e epidermis-specific knockout mice (\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003efl/fl\u003c/em\u003e\u003c/sup\u003e-\u003cem\u003eK14-Cre\u003c/em\u003e\u003csup\u003e\u003cem\u003eERT\u003c/em\u003e\u003c/sup\u003e). When \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003efl/fl\u003c/em\u003e\u003c/sup\u003e-\u003cem\u003eK14-Cre\u003c/em\u003e\u003csup\u003e\u003cem\u003eER\u003c/em\u003e\u003c/sup\u003e was induced by tamoxifen for 6 weeks, we called them \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e mice. \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003efl/fl\u003c/em\u003e\u003c/sup\u003e-\u003cem\u003eK14\u003c/em\u003e-\u003cem\u003eCre\u003c/em\u003e\u003csup\u003e\u003cem\u003eERT\u003c/em\u003e\u003c/sup\u003e were crossed to \u003cem\u003eZbtb46-DTR\u003c/em\u003e mice to generate DC knockout in \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e mice (\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e-\u003cem\u003eZbtb\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u0026minus;/\u0026minus;\u003c/em\u003e\u003c/sup\u003e). \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003efl/fl\u003c/em\u003e\u003c/sup\u003e-\u003cem\u003eK14\u003c/em\u003e-\u003cem\u003eCre\u003c/em\u003e\u003csup\u003e\u003cem\u003eERT\u003c/em\u003e\u003c/sup\u003e were crossed to \u003cem\u003eRag1\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u0026minus;/\u0026minus;\u003c/em\u003e\u003c/sup\u003e mice (GemPharmatech) to generate DC knockout in \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e\u003cem\u003e△Epi\u003c/em\u003e\u003c/sup\u003e mice (\u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003e△Epi\u003c/sup\u003e-\u003cem\u003eRag1\u003c/em\u003e\u003csup\u003e\u003cem\u003e\u0026minus;/\u0026minus;\u003c/em\u003e\u003c/sup\u003e). Sex- and age-matched animals between 12 and 15 weeks of age were used for experiments. Preliminary experiments were performed to determine proper sample size. All animal experiments were performed in accordance with protocols approved by the Second Affiliated Hospital, Zhejiang University School of Medicine Animal Care Committee (2019-No.072).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec11\" class=\"Section2\"\u003e \u003ch2\u003eFlow cytometric analysis of skin tissue-derived cells\u003c/h2\u003e \u003cp\u003eMouse/human skin biopsy tissue was immersed in 0.5% dispase (GibcoTM, USA) at 4\u0026deg;C overnight in order to separate the epidermis and dermis. The epidermis was digested with 0.25% trypsin (Thermo Fisher Scientific, USA), while the dermis was digested with 1 mg/ml collagenase (Sigma-Aldrich, USA). After neutralization with serum, the mixture was centrifuged at 1000 rpm for 5 min, and the cells were resuspended in PBS to obtain a single-cell suspension.\u003c/p\u003e \u003cp\u003eEpidermal and dermal single-cell suspensions were prepared as described above. The cells were first stimulated with Cell Activation Cocktail (with Brefeldin A, BioLegend) together with 2 \u0026micro;g/ml resiquimod (Sigma) for 6 hours. After stimulation, the cells were first stained with a Zombie UV Fixable Viability Kit (BioLegend) for 15 minutes and then incubated with the Fc receptor blocker TruStain FcX (BioLegend) for 10 minutes. Then, the cells were incubated with mixed cell surface antibodies for 30 minutes at 4\u0026deg;C in the dark. For intracellular staining, cells were fixed and permeabilized by using the BD Cytofix/Cytoperm Kit (BD Biosciences) or Foxp3/Transcription Factor Staining Buffer Set (eBioscience) and then incubated with anti-cytokine antibodies for 30 minutes at 4\u0026deg;C in the dark. The antibodies used were purchased from BioLegend, BD Biosciences, or eBioscience, as listed in supplementary methods. The data were acquired by CytoFLEX LX (Beckman Coulter) and analyzed by CytExpert Software (Beckman Coulter).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec12\" class=\"Section2\"\u003e \u003ch2\u003eSingle-cell RNA sequencing data analysis\u003c/h2\u003e \u003cp\u003eProcessed human single-cell RNA-sequencing (scRNA-seq) datasets from our datasets, sequenced by performed by PLTTech Inc. (Hangzhou, China). For human skin integration of the psoriatic epidermis (PE) and normal epidermis (NE) was analyzed with the Seurat R package (versions 3.0.0 and 4.0.3) \u003csup\u003e\u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e48\u003c/span\u003e\u003c/sup\u003e. Cells with \u0026gt;\u0026thinsp;20% mitochondrial gene percentage, minimum of 350 genes and maximum of 4500 detected were filtered from downstream analyses. Samples were split by donor identity (referred to as sample in metadata in analysis scripts and Seurat objects) into individual objects by donor, which were each normalized and processed with the NormalizeData and FindVariableFeatures functions with default parameters. Integration anchors were determined with these donor objects using the first 30 dimensions and integrated using the IntegrateData function on the first 30 dimensions. Using this integrated object, dimensionality reduction was carried out after ScaleData with principal component analysis and Uniform manifold approximation and projection (UMAP) on the first 30 principal components to generate UMAP. T lymphocytes (Human NE, Human PE) clustering was performed at resolution 0.2 after FindNeighbors was run on the first 30 dimensions. Myeloid cells clustering was performed at resolution 0.5 after FnidNeighbors was run on the first 30 dimensions.\u003c/p\u003e \u003cp\u003eSimilarly, for mouse skin integrated data, a Seurat object was created with 10x cellranger outputs. SingleCell Data Analysis was performed using the OmicStudio tools created by LC-BIO Co., Ltd (HangZhou, China) at \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://www.omicstudio.cn/cell\u003c/span\u003e\u003cspan address=\"https://www.omicstudio.cn/cell\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e, and was analyzed with the Seurat R package (4.1.0, R version 4.1.3).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec13\" class=\"Section2\"\u003e \u003ch2\u003eAnti-IL-17A/ Vγ2/ CXCL16 treatment\u003c/h2\u003e \u003cp\u003eThe procedure of anti-IL-17A treatment was attached in Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003el with 50ug/g/week hamster anti-mouse mAb (17F3) by subcutaneous injection (s.c.) and respective hamster IgG isotype control antibody (BE0091, both Bio X Cell) (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ej). Anti-Vγ2 treatment was performed by intraperitoneal injection (i.p.) of the mice with 10 \u0026micro;g/g/week hamster anti-mouse mAb (UC3-10A6) and respective hamster IgG isotype control antibody (BE0091, both Bio X Cell) (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ej). Anti-mouse CXCL16 treatment (R\u0026amp;D, AF503), was performed by subcutaneous injecti4on (s.c.) of the mice with 4ug/g/week and respective hamster IgG isotype control antibody (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003eh).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec14\" class=\"Section2\"\u003e \u003ch2\u003ePrimary keratinocyte culture\u003c/h2\u003e \u003cp\u003eThe separated epidermis was digested with 0.25% trypsin (Thermo Fisher Scientific, USA). The digested epidermal keratinocytes were neutralized with FBS (Gibco, USA) and centrifuged at 1000 rpm. Cells were cultured to 70% confluence in EpiGRO\u0026trade; Human Epidermal Keratinocyte Complete Culture Media Kit (Millipore, USA, SCMK001) in a humidified incubator with 5% CO\u003csub\u003e2\u003c/sub\u003e at 37\u0026deg;C.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec15\" class=\"Section2\"\u003e \u003ch2\u003eRecombinant mouse IL-23/CXCL16 treatment\u003c/h2\u003e \u003cp\u003eMice were injected intradermally (i.d.) with 20 \u0026micro;L PBS/0.1% bovine serum albumin (BSA) containing 0.5 \u0026micro;g recombinant mouse (rm)IL-23 or recombinant mouse (rm)CXCL16 in the right ear using a 29-gauge needle, every other day for 14 days.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec16\" class=\"Section2\"\u003e \u003ch2\u003eQuantification and statistical analysis\u003c/h2\u003e \u003cp\u003eStatistical analyses were performed with GraphPad Prism, versions 9\u0026ndash;10 (GraphPad Software, San Diego, CA), or R, versions 3.5.1, 3.61, and 4.0.2. Parameters such as number of replicates; the number of independent experiments; measures of center, dispersion, and precision (mean SD or SEM); statistical test; and significance are reported in figures and figure legends. P values are indicated as follows: \u0026lowast; P\u0026thinsp;\u0026lt;\u0026thinsp;0.05, \u0026lowast;\u0026lowast; P\u0026thinsp;\u0026lt;\u0026thinsp;0.01, \u0026lowast;\u0026lowast;\u0026lowast; P\u0026thinsp;\u0026lt;\u0026thinsp;0.001.\u003c/p\u003e \u003c/div\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eFunding\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study was supported by grants from the National Natural Science Foundation of China (No. 82230104, 81930089, 81773318, 82404117, 82103709 and 82303999).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor Contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eConceptualization: XYC, LRY; Formal Analysis: XYC, XYM; Funding Acquisition: XYM; Investigation: XYC, LRY, XYM, NCF, XBC; Methodology: BXY, NCF, YZC, XYM, XYC, SQC; YXZ, MYL; Project Administration: XYM; Resources: XYM; Supervision: MZ, XYM; Validation: XYC, YZ, NCF; Writing and original Draft Preparation: XYC, LRY; Writing - Review and Editing: LRY, XYC, XYM.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare no competing interests.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData and materials availability:\u0026nbsp;\u003c/strong\u003eAll data needed to evaluate the conclusions in the paper are present in the paper or the Supplementary Materials. The bulk-RNA-seq data and scRNA-seq data are available in the Gene Expression Omnibus (GEO) repository database and are submitting. The \u003cem\u003eUbe2l3\u003c/em\u003e\u003csup\u003efl/fl\u0026nbsp;\u003c/sup\u003emice developed for this study will be shared upon request. All data needed to evaluate the conclusions in the paper are present in the paper or the Supplementary Materials.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eC. E. M. Griffiths, A. W. Armstrong, J. E. Gudjonsson, J. Barker, Psoriasis. \u003cem\u003eLancet\u003c/em\u003e \u003cstrong\u003e397\u003c/strong\u003e, 1301-1315 (2021).\u003c/li\u003e\n\u003cli\u003eW. H. Boehncke, M. P. Schon, Psoriasis. \u003cem\u003eLancet\u003c/em\u003e \u003cstrong\u003e386\u003c/strong\u003e, 983-994 (2015).\u003c/li\u003e\n\u003cli\u003e\u0026Ouml;. Ulu\u0026ccedil;kan, E. F. 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Stuart\u003cem\u003e et al.\u003c/em\u003e, Comprehensive Integration of Single-Cell Data. \u003cem\u003eCell\u003c/em\u003e\u003cstrong\u003e177\u003c/strong\u003e, 1888-1902 e1821 (2019).\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"nature-portfolio","isNatureJournal":true,"hasQc":false,"allowDirectSubmit":false,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"","title":"Nature Portfolio","twitterHandle":"","acdcEnabled":false,"dfaEnabled":false,"editorialSystem":"ejp","reportingPortfolio":"","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"","lastPublishedDoi":"10.21203/rs.3.rs-5973089/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-5973089/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003ePsoriasis is a chronic, complex immune-mediated inflammatory disorder with cutaneous and systemic manifestations in which keratinocytes, dendritic cells and T cells have central roles. UBE2L3 may be a protective biomarker that regulates the pathogenesis of psoriasis. Here, we identified the IL-17A signaling similarity between human psoriatic skin and \u003cem\u003eUbe2l3\u003c/em\u003e conditional knockout mouse skin in the epidermis rather than dermis. IL-17A was regulated by CXCR6\u003csup\u003e+\u003c/sup\u003e Vγ2\u003csup\u003e+ \u003c/sup\u003eγδT in mouse while CXCR6\u003csup\u003e+ \u003c/sup\u003eCD8\u003csup\u003e+ \u003c/sup\u003eT in human. CXCL16 is the only chemokines whose bind to stimulate CXCR6. \u003cem\u003eUbe2l3\u003c/em\u003e reduction in keratinocytes activated IL-1β and then promote CXCL16 expression through STAT3 signaling. Up-regulated CXCL16 in keratinocytes and cDC2/mDC then attracted Vγ2\u003csup\u003e+ \u003c/sup\u003eγδT17 or CD8\u003csup\u003e+ \u003c/sup\u003eT to secrete IL-17A and form a positive feedback loop in keratinocytes supporting psoriatic lesion. Thus, UBE2L3 is a keratinocyte-intrinsic suppressor of epidermal IL-17 production in Vγ2\u003csup\u003e+ \u003c/sup\u003eγδT in mouse and CD8\u003csup\u003e+ \u003c/sup\u003eT in human through CXCL16/CXCR6 signaling pathway in psoriasis.\u003c/p\u003e","manuscriptTitle":"Cell atlas of human psoriasis and epidermal specific Ube2l3 deficiency mouse highlighting CXCL16/CXCR6 orchestrating the development of psoriasis","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-02-28 13:31:50","doi":"10.21203/rs.3.rs-5973089/v1","editorialEvents":[],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"nature-communications","isNatureJournal":true,"hasQc":false,"allowDirectSubmit":false,"externalIdentity":"NCOMMS","sideBox":"Learn more about [Nature Communications](http://www.nature.com/ncomms/)","snPcode":"","submissionUrl":"https://mts-ncomms.nature.com/","title":"Nature Communications","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"ejp","reportingPortfolio":"Nature Communications","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"c2e69796-59b0-4138-bef5-3b2740eb2f97","owner":[],"postedDate":"February 28th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[{"id":44896018,"name":"Biological sciences/Immunology/Inflammation/Chronic inflammation"},{"id":44896019,"name":"Health sciences/Biomarkers/Prognostic markers"},{"id":44896020,"name":"Health sciences/Molecular medicine"}],"tags":[],"updatedAt":"2025-10-14T07:08:04+00:00","versionOfRecord":{"articleIdentity":"rs-5973089","link":"https://doi.org/10.1038/s41467-025-64106-6","journal":{"identity":"nature-communications","isVorOnly":false,"title":"Nature Communications"},"publishedOn":"2025-10-13 04:00:00","publishedOnDateReadable":"October 13th, 2025"},"versionCreatedAt":"2025-02-28 13:31:50","video":"","vorDoi":"10.1038/s41467-025-64106-6","vorDoiUrl":"https://doi.org/10.1038/s41467-025-64106-6","workflowStages":[]},"version":"v1","identity":"rs-5973089","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-5973089","identity":"rs-5973089","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}