cGAS-STING dependent type I IFN protects against Leptospira interrogans renal colonization in mice

cGAS-STING dependent type I IFN protects against Leptospira interrogans renal colonization in mice | bioRxiv /* */ /* */ <!-- <!-- /*! * yepnope1.5.4 * (c) WTFPL, GPLv2 */ (function(a,b,c){function d(a){return"[object Function]"==o.call(a)}function e(a){return"string"==typeof a}function f(){}function g(a){return!a||"loaded"==a||"complete"==a||"uninitialized"==a}function h(){var a=p.shift();q=1,a?a.t?m(function(){("c"==a.t?B.injectCss:B.injectJs)(a.s,0,a.a,a.x,a.e,1)},0):(a(),h()):q=0}function i(a,c,d,e,f,i,j){function k(b){if(!o&&g(l.readyState)&&(u.r=o=1,!q&&h(),l.onload=l.onreadystatechange=null,b)){"img"!=a&&m(function(){t.removeChild(l)},50);for(var d in y[c])y[c].hasOwnProperty(d)&&y[c][d].onload()}}var j=j||B.errorTimeout,l=b.createElement(a),o=0,r=0,u={t:d,s:c,e:f,a:i,x:j};1===y[c]&&(r=1,y[c]=[]),"object"==a?l.data=c:(l.src=c,l.type=a),l.width=l.height="0",l.onerror=l.onload=l.onreadystatechange=function(){k.call(this,r)},p.splice(e,0,u),"img"!=a&&(r||2===y[c]?(t.insertBefore(l,s?null:n),m(k,j)):y[c].push(l))}function j(a,b,c,d,f){return q=0,b=b||"j",e(a)?i("c"==b?v:u,a,b,this.i++,c,d,f):(p.splice(this.i++,0,a),1==p.length&&h()),this}function k(){var a=B;return a.loader={load:j,i:0},a}var l=b.documentElement,m=a.setTimeout,n=b.getElementsByTagName("script")[0],o={}.toString,p=[],q=0,r="MozAppearance"in l.style,s=r&&!!b.createRange().compareNode,t=s?l:n.parentNode,l=a.opera&&"[object Opera]"==o.call(a.opera),l=!!b.attachEvent&&!l,u=r?"object":l?"script":"img",v=l?"script":u,w=Array.isArray||function(a){return"[object Array]"==o.call(a)},x=[],y={},z={timeout:function(a,b){return b.length&&(a.timeout=b[0]),a}},A,B;B=function(a){function b(a){var a=a.split("!"),b=x.length,c=a.pop(),d=a.length,c={url:c,origUrl:c,prefixes:a},e,f,g;for(f=0;f<d;f++)g=a[f].split("="),(e=z[g.shift()])&&(c=e(c,g));for(f=0;f<b;f++)c=x[f](c);return c}function g(a,e,f,g,h){var i=b(a),j=i.autoCallback;i.url.split(".").pop().split("?").shift(),i.bypass||(e&&(e=d(e)?e:e[a]||e[g]||e[a.split("/").pop().split("?")[0]]),i.instead?i.instead(a,e,f,g,h):(y[i.url]?i.noexec=!0:y[i.url]=1,f.load(i.url,i.forceCSS||!i.forceJS&&"css"==i.url.split(".").pop().split("?").shift()?"c":c,i.noexec,i.attrs,i.timeout),(d(e)||d(j))&&f.load(function(){k(),e&&e(i.origUrl,h,g),j&&j(i.origUrl,h,g),y[i.url]=2})))}function h(a,b){function c(a,c){if(a){if(e(a))c||(j=function(){var a=[].slice.call(arguments);k.apply(this,a),l()}),g(a,j,b,0,h);else if(Object(a)===a)for(n in m=function(){var b=0,c;for(c in a)a.hasOwnProperty(c)&&b++;return b}(),a)a.hasOwnProperty(n)&&(!c&&!--m&&(d(j)?j=function(){var a=[].slice.call(arguments);k.apply(this,a),l()}:j[n]=function(a){return function(){var b=[].slice.call(arguments);a&&a.apply(this,b),l()}}(k[n])),g(a[n],j,b,n,h))}else!c&&l()}var h=!!a.test,i=a.load||a.both,j=a.callback||f,k=j,l=a.complete||f,m,n;c(h?a.yep:a.nope,!!i),i&&c(i)}var i,j,l=this.yepnope.loader;if(e(a))g(a,0,l,0);else if(w(a))for(i=0;i (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];var j=d.createElement(s);var dl=l!='dataLayer'?'&l='+l:'';j.src='//www.googletagmanager.com/gtm.js?id='+i+dl;j.type='text/javascript';j.async=true;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-M677548'); Skip to main content Home About Submit ALERTS / RSS Search for this keyword Advanced Search New Results cGAS-STING dependent type I IFN protects against Leptospira interrogans renal colonization in mice Suman Gupta , James Matsunaga , Bridget Ratitong , Sana Ismaeel , Diogo G. Valadares , View ORCID Profile A. Phillip West , Nagaraj Kerur , Christian Stehlik , Andrea Dorfleutner , Jargalsaikhan Dagvadorj , Jenifer Coburn , Andrea J. Wolf , Suzanne L. Cassel , David A. Haake , View ORCID Profile Fayyaz S. Sutterwala doi: https://doi.org/10.1101/2025.06.02.657349 Suman Gupta 1 Department of Medicine, Cedars-Sinai Medical Center , Los Angeles, California, USA 2 Women’s Guild Lung Institute, Cedars-Sinai Medical Center , Los Angeles, California, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site James Matsunaga 3 Research Service, VA Greater Los Angeles Healthcare System , Los Angeles, California, USA 4 Department of Medicine, David Geffen School of Medicine, University of California , Los Angeles, California, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Bridget Ratitong 1 Department of Medicine, Cedars-Sinai Medical Center , Los Angeles, California, USA 2 Women’s Guild Lung Institute, Cedars-Sinai Medical Center , Los Angeles, California, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Sana Ismaeel 5 Department of Pathology and Laboratory Medicine, Cedars-Sinai Medical Center , Los Angeles, California, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Diogo G. Valadares 1 Department of Medicine, Cedars-Sinai Medical Center , Los Angeles, California, USA 2 Women’s Guild Lung Institute, Cedars-Sinai Medical Center , Los Angeles, California, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site A. Phillip West 6 The Jackson Laboratory , Bar Harbor, Maine, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for A. Phillip West Nagaraj Kerur 7 Department of Ophthalmology and Visual Sciences, The Ohio State University Wexner Medical Center , Columbus, Ohio, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Christian Stehlik 5 Department of Pathology and Laboratory Medicine, Cedars-Sinai Medical Center , Los Angeles, California, USA 8 Department of Biomedical Sciences, Cedars-Sinai Medical Center , Los Angeles, California, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Andrea Dorfleutner 5 Department of Pathology and Laboratory Medicine, Cedars-Sinai Medical Center , Los Angeles, California, USA 8 Department of Biomedical Sciences, Cedars-Sinai Medical Center , Los Angeles, California, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Jargalsaikhan Dagvadorj 1 Department of Medicine, Cedars-Sinai Medical Center , Los Angeles, California, USA 2 Women’s Guild Lung Institute, Cedars-Sinai Medical Center , Los Angeles, California, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Jenifer Coburn 9 Department of Microbiology and Immunology, Medical College of Wisconsin , Milwaukee, Wisconsin, USA 10 Department of Medicine, Medical College of Wisconsin , Milwaukee, Wisconsin, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Andrea J. Wolf 8 Department of Biomedical Sciences, Cedars-Sinai Medical Center , Los Angeles, California, USA 11 F. Widjaja Inflammatory Bowel Disease Institute, Cedars-Sinai Medical Center , Los Angeles, California, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Suzanne L. Cassel 1 Department of Medicine, Cedars-Sinai Medical Center , Los Angeles, California, USA 2 Women’s Guild Lung Institute, Cedars-Sinai Medical Center , Los Angeles, California, USA 8 Department of Biomedical Sciences, Cedars-Sinai Medical Center , Los Angeles, California, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site For correspondence: suzanne.cassel{at}cshs.org DHaake{at}UCLA.edu fayyaz.sutterwala{at}cshs.org David A. Haake 12 Division of Infectious Diseases , VA Greater Los Angeles Healthcare System, Los Angeles, California, USA 13 Department of Medicine, David Geffen School of Medicine, University of California , Los Angeles, California, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site For correspondence: suzanne.cassel{at}cshs.org DHaake{at}UCLA.edu fayyaz.sutterwala{at}cshs.org Fayyaz S. Sutterwala 1 Department of Medicine, Cedars-Sinai Medical Center , Los Angeles, California, USA 2 Women’s Guild Lung Institute, Cedars-Sinai Medical Center , Los Angeles, California, USA 8 Department of Biomedical Sciences, Cedars-Sinai Medical Center , Los Angeles, California, USA 13 Department of Medicine, David Geffen School of Medicine, University of California , Los Angeles, California, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Fayyaz S. Sutterwala For correspondence: suzanne.cassel{at}cshs.org DHaake{at}UCLA.edu fayyaz.sutterwala{at}cshs.org Abstract Full Text Info/History Metrics Preview PDF Abstract Leptospira interrogans is the major causative agent of leptospirosis. Humans, canines and livestock animals are susceptible to Leptospira species and can develop fulminant disease. Rodents serve as reservoir hosts in which the bacteria colonize the renal tubules and are excreted in the urine. The host immune response to Leptospira spp. remains poorly defined. We show that L. interrogans induces a robust type I interferon (IFN) response in human and murine macrophages that is dependent on the cytosolic dsDNA sensor Cyclic GMP-AMP Synthase (cGAS) and the Stimulator of IFN Genes (STING) signaling pathway. Further, we show that mice deficient in the IFNα/β receptor subunit 1 (IFNAR1) or STING had higher bacterial burdens and increased renal colonization following infection in vivo suggesting that cGAS-STING-driven type I IFN is required for the host defense against L. interrogans . These findings demonstrate the significance of cGAS-STING-dependent type I IFN signaling in mammalian innate immune responses to L. interrogans . Author Summary Leptospirosis is a globally distributed zoonotic disease caused by spirochetes belonging to the genus Leptospira . While humans, livestock, and dogs can develop severe or even fatal illness upon infection, rodents typically serve as asymptomatic reservoir hosts. A defining feature of the leptospiral life cycle is the ability of the pathogen to colonize the kidney in these reservoir host species, leading to prolonged urinary shedding and environmental dissemination. Despite the significant global burden of leptospirosis, the innate immune pathways that detect this pathogen and prevent renal colonization remain poorly understood. In this study we demonstrate that L. interrogans induces a robust type I IFN cytokine response from macrophages. The induction of this type I IFN response is dependent on sensing cytosolic DNA by the cGAS-STING pathway. Using in vivo mouse models of L. interrogans infection we further show that activation of this pathway is required to control bacterial burdens and reduce long-term kidney colonization. This study is the first to demonstrate a critical role for cGAS-STING and type I IFN in controlling L. interrogans infection. Introduction Leptospirosis is caused by didermal spirochetes belonging to the genus Leptospira with L. interrogans being responsible for most pathogenic cases ( 1 ). All vertebrates are susceptible to infection, and the manifestation of symptoms varies extensively in different hosts ( 2 ). Cattle and canines experience mild to severe illnesses, indicating a heightened sensitivity to leptospirosis. In contrast, rodents such as mice and rats typically remain asymptomatic while being chronically colonized in their kidneys, serving as reservoir hosts for Leptospira spp. and shedding the bacteria in their urine for long periods of time ( 3 – 5 ). This process leads to soil and water contamination, perpetuating the enzootic cycle of the disease. Humans are incidental hosts with many infections remaining asymptomatic. However, humans can develop multiorgan dysfunction which can prove fatal in 10-20% of cases ( 6 ). Leptospires are stealth pathogens that evade recognition by components of the host innate immune system. Leptospires have been shown to evade TLR5 activation due to the periplasmic positioning of their endoflagella ( 7 ). Additionally, they avoid detection by NOD1/NOD2 by tightly associating peptidoglycan with the outer membrane lipoprotein LipL21 which prevents the release of immunologically active peptidoglycan fragments ( 8 ). Despite these immune evasion strategies, leptospires also stimulate innate immune responses via other pathways. In both human and murine cells, leptospires are potent agonists of TLR2, primarily through their tri-acylated lipoproteins ( 9 , 10 ). Leptospira possess lipopolysaccharide (LPS) in their outer membrane, which serve as a virulence factor ( 11 ). Leptospiral LPS is also known to be recognized by TLR4 in mice ( 12 ). TLR2 and TLR4 recognition of leptospires has been identified as critical for murine resistance to acute leptospirosis ( 13 ). The role of macrophages in the detection and clearance of leptospires remains incompletely understood. Depletion of peritoneal macrophages in C57BL/6J mice was shown to increase sensitivity to leptospirosis, resulting in an increased bacterial burden ( 14 – 17 ). In addition, macrophage depletion led to increased kidney colonization ( 16 , 17 ). Leptospires were generally classified as extracellular bacteria, although more recent studies have suggested the presence of leptospires within macrophages ( 18 – 20 ). Leptospires may either exist freely in the cytosol or reside within specific cellular compartments, such as endosomes, lysosomes, or phagosomes ( 19 ). The fate of internalized leptospires is unclear, but recent findings indicate that leptospires do not replicate within mouse macrophages and that viable internalized leptospires can exit these cells over time ( 21 ). The precise mechanisms by which macrophages detect leptospires and contribute to their control remains inadequately understood. Intracellular innate immune pathways that sense the presence of cytosolic nucleic acids can play a critical role in the generation of type I interferon (IFN) responses. Cyclic GMP-AMP synthase (cGAS) is one such intracellular sensor that can bind to double-stranded DNA in the cytosol ( 22 – 26 ). Upon binding, cGAS generates 2’3’-cyclic guanosine monophosphate-adenosine monophosphate (cGAMP), which activates stimulator of interferon genes (STING). This activation results in the recruitment and subsequent phosphorylation of Tank-binding kinase-1 (TBK1) and IFN regulatory factor 3 (IRF3), followed by the production and secretion of IFNa and IFNý. Type I IFN signaling through the IFNα/β receptor (IFNAR) induces the transcription of interferon-stimulated genes (ISGs) ( 27 , 28 ). The cGAS-STING pathway was initially shown to play a role in anti-viral host response; it has since been shown to have roles in immune responses to a variety of bacterial pathogens including Listeria monocytogenes, Mycobacterium tuberculosis, Klebsiella pneumoniae , and Borrelia burgdorferi ( 23 , 25 , 26 , 29 ). Bacteria that produce cyclic dinucleotides can directly activate STING independently from cGAS ( 30 – 33 ). Whether Leptospira can activate the cGAS-STING pathway and the physiologic relevance of type I IFN production in leptospirosis is currently unknown. Because pathogenic leptospires possess genes involved in cyclic dinucleotide synthesis ( 34 , 35 ) and leptospires can be internalized by macrophages, we asked if cGAS-STING-mediated type I IFN signaling was involved in the host immune responses against leptospiral infection. In this study, we demonstrate that L. interrogans induces type I IFN production and the transcriptionof ISGs in both mouse and human macrophages. The type I IFN response was found to be dependent on effective phagocytosis and degradation of leptospires in the macrophages. Further, we show that the type I IFN induction in L. interrogans -infected macrophages involves the cGAS-STING pathway. Finally, the absence of IFNAR or STING in vivo resulted in increased susceptibility to L. interrogans infection and increased kidney colonization, highlighting the importance of cGAS-STING-driven type I IFN in mammalian host defense against L. interrogans . Results Leptospira induces a type I IFN response following infection of macrophages We first evaluated the ability of mouse macrophages to induce a type I IFN response following infection with L. interrogans . Consistent with prior reports we observed that bone marrow-derived macrophages (BMDMs) infected with L. interrogans serovar Copenhageni strain Fiocruz L1-130 at a multiplicity of infection (MOI) of 100 elicited the secretion of TNF, IL-10, and IL-6 ( Fig 1A-C ) ( 13 , 36 ). Additionally, robust secretion of IFNβ was also observed ( Fig 1D and S1A ). Tank-binding kinase 1 (TBK1) and IFN regulatory factor 3 (IRF3) play central roles in the initiation of type I IFN responses following signaling by the mitochondrial antiviral signaling protein (MAVS), STING, and TICAM1 ( 37 ). We found L . interrogans infection of BMDM resulted in the phosphorylation of TBK1 and IRF3 consistent with the observed production of IFNβ ( Fig 1E ). We also assessed the transcript levels of Ifna and Ifnb in BMDMs and observed upregulation of both transcripts by 4 hours post-infection ( Fig 1F, G ). Similar kinetics for IFN-β transcript upregulation were observed in PMA-differentiated human THP-1 cells ( Fig 1H ) suggesting L. interrogans can induce a type I IFN response in both mouse and human macrophages. Download figure Open in new tab Fig 1. L. interrogans infection of macrophages induces a type I IFN response. (A-D) Quantification of TNF, IL-10, IL-6, and IFNβ by ELISA in culture supernatants of WT BMDMs infected with L. interrogans Fiocruz L1-130 (MOI 100) for 4, 6, and 8h. (E) Immunoblot for p-TBK1, total TBK1, p-IRF3, total IRF3 and β-actin in cell lysates of BMDMs infected with L. interrogans (MOI 100) for 2, 4, and 6h. Data are representative of three independent experiments. (F, G) Relative expression of Ifna and Ifnb transcript levels in BMDMs infected with L. interrogans at 4 and 6h post infection. (H) Relative expression of IFNB transcript levels in PMA-differentiated human THP-1 cells infected with L. interrogans at 4, 6, and 8h post infection. (A-D and F-H) Data are pooled from three independent experiments and expressed as the mean ± SEM. Phagocytic uptake of L. interrogans is required for effective IFNβ secretion. To determine if L. interrogans were being phagocytosed, BMDMs were infected with pHrodo-labelled bacteria and increase in fluorescence dependent on phagosomal acidification was assessed. We found a robust increase in florescence of pHrodo-labelled bacteria with time suggesting bacteria were being taken up in acidic phagosomal compartment ( Fig 2A ). Additionally, wild-type (WT) BMDMs pretreated with cytochalasin D, an inhibitor of actin polymerization, prior to infection with pHrodo-labelled L. interrogans showed a reduction in fluorescence, suggesting that the uptake of bacteria into acidic subcellular compartments is via a phagocytic pathway ( Fig 2A ). After phagocytosis, bacteria-containing phagosomes recruit Rab5 to their surface, which is then replaced by LAMP1 upon lysosomal fusion ( 38 – 41 ). We found a subset of internalized bacteria colocalizing with Rab5 at 3 hours post infection ( Fig 2B ). Some bacteria were also found to be colocalizing with LAMP1 ( Fig 2B ) consistent with phagocytic uptake and processing of bacteria. Download figure Open in new tab Fig 2. Phagocytosis of L. interrogans is required for macrophage driven IFNβ production. (A) Increase in mean fluorescence intensity per cell (Red Calibrated Unit; RCU) in WT BMDMs, after addition of L. interrogans Fiocruz L1-130 (MOI 100) labelled with pH-sensitive dye pHrodo, read by Incucyte SX5 reader at 1h intervals for a period of 12h in the presence or absence of cytochalasin D (2µM). Curves are representative of three independent experiments. Area under the curve (AUC) was calculated for the increase in pHrodo fluorescence intensity per cell in WT BMDMs in the presence or absence of cytochalasin D. Data are pooled from three independent experiments and expressed as the mean ± SEM. (B) Representative immunofluorescence image showing co-localization (arrow heads) of L. interrogans Fiocruz L1-130 with Rab5 and LAMP1 in BMDM at 3h post infection. Scale bar = 10 μm. Quantification of co-localization by pixel-pixel overlap calculated by JaCoP plugin in ImageJ. Data are pooled from three frames per experiment from three independent experiments. (C-E) Quantification of IFNβ and TNF by ELISA in culture supernatants of WT BMDMs pretreated with cytochalasin D, bafilomycin A1, and chymostatin, followed by infection with L. interrogans (MOI 100) for 6h. Data pooled from three experiments and expressed as the mean ± SEM. Statistical significance was calculated by two-way ANOVA; *p<0.05, ***p<0.001, ns=non-significant. To determine if phagocytosis is required for the production of IFNβ, BMDMs were pretreated with cytochalasin D prior to infection with L. interrogans . Cytochalasin D treatment resulted in diminished IFNβ secretion ( Fig 2C ). Furthermore, treatment with bafilomycin A1, which inhibits phagosome and lysosomal acidification by blocking the vacuolar ATPase pump ( 42 – 44 ), and chymostatin, an inhibitor of phagosomal proteases ( 45 , 46 ), each resulted in reduced IFNβ following infection with L. interrogans ( Fig 2D ). Cytochalasin D, bafilomycin A1, and chymostatin did not affect TNF secretion from BMDM in response to infection with L. interrogans ( Fig 2E ). These findings suggest that L. interrogans phagocytosis by macrophages is crucial for a robust type I IFN response. Challenge of BMDM with heat-killed L. interrogans still resulted in IFNβ secretion indicating that live bacteria were not required for induction of the IFNβ response ( S1B Fig ). The induction of IFNβ secretion was also not dependent on the pathogenicity of the bacteria, as both pathogenic L. interrogans Fiocruz L1-130 and saprophytic L. biflexa serovar Patoc strain Patoc 1 strains induced similar levels of IFNβ ( S1C Fig ). Type I IFN signaling is required for effective control of L. interrogans in mice in vivo Given that Leptospira induced a robust type I IFN response, we next assessed if the induction of type I IFN was important for the host’s ability to control bacterial replication in a mouse model of infection. C57BL/6J mice infected with L. interrogans do not develop fulminant disease; however, L. interrogans does chronically colonize the kidneys where they can be shed in the urine ( 47 – 49 ). To investigate the role of type I IFN signaling in acute L. interrogans infection, we used mice deficient in the IFNα/β receptor subunit 1 (IFNAR1). As expected, in vitro analysis of BMDMs from Ifnar1 -/- and WT mice showed no difference in IFNβ secretion following infection with L. interrogans ( Fig 3A ); however, the expression of ISGs, such as Ifit1, Ifit3, Ifi44 , and Zbp1 was significantly diminished in L. interrogans infected Ifnar1 -/- BMDMs compared to WT ( Fig 3B ). Download figure Open in new tab Fig 3. Increased susceptibility of IFNAR-deficient mice to L. interrogans in vivo . (A) Quantification of IFNβ by ELISA in culture supernatants of WT and Ifnar1 -/- BMDMs infected with L. interrogans Fiocruz L1-130 (MOI 100) for 4, 6, and 8h. Data are pooled from three experiments and expressed as the mean ± SEM. Statistical significance was calculated by two-way ANOVA. (B) Relative expression of ifit1, ifit3, ifi44, and zbp1 transcript levels in WT and Ifnar1 -/- BMDMs infected with L. interrogans at 6h post infection. Data pooled from three experiments and expressed as the mean ± SEM. (C) Schematic of acute infection model; WT and Ifnar1 -/- mice were infected intraperitoneally with 2×10 8 CFU of L. interrogans Fiocruz L1-130. (D, E) Bacterial abundance in the spleen and kidney at day 3 post-infection was assessed by qPCR for the L. interrogans lipL32 gene and normalized to the eukaryotic PPIA gene. Data are pooled from two independent experiments: n=9 WT, n=10 Ifnar1 -/- . (F) Relative expression of Ifna and Ifnb transcripts in spleen 24h post-acute infection in WT and Ifnar1 -/- mice. n=5 WT, n=5 Ifnar1 -/- . (G) Schematic of chronic infection model; WT and Ifnar1 -/- mice were infected subcutaneously with 1×10 6 CFU of L. interrogans Fiocruz L1-130. (H) Bacterial abundance in the kidney at day 14 post-infection was assessed by qPCR as above. Data are pooled from two independent experiments: n=6 WT, n=6 Ifnar1 -/- . (D, E, H) Statistical significance was calculated by Mann-Whitney U test. (F) Statistical significance calculated by two-way ANOVA; *p<0.05, **p<0.01, ****p<0.0001, ns=non-significant. Following an intraperitoneal in vivo infection with 2×10 8 L. interrogans ( Fig 3C ), we found a higher bacterial burden in the spleen of Ifnar1 -/- mice compared to WT at 3 days post-infection ( Fig 3D ). In contrast, no difference in bacterial load was observed in the kidneys at 3 days post-infection ( Fig 3E ). As expected, transcript analysis of Ifna and Ifnb in the spleen at 24 hours post-infection revealed no differences between Ifnar1 -/- mice and WT mice ( Fig 3F ). Together these data demonstrate that L. interrogans induces the transcription of ISGs in L. interrogans infected macrophages and that type I IFN signaling through IFNAR1 is important for bacterial control in an acute leptospiral infection. To assess renal colonization by L. interrogans , mice were infected with 1×10 6 spirochetes subcutaneously and kidney colonization assessed at 14 days post-infection ( Fig 3G ). Ifnar1 -/- mice had higher kidney colonization with L. interrogans compared to WT mice ( Fig 3H ). Taken together these data suggest that type I IFN signaling through IFNAR plays an important role in vivo in acutely controlling L. interrogans replication and limiting chronic kidney colonization. L. interrogans -induced type I IFN is cGAS-STING dependent As the type I IFN response from BMDMs in response to L. interrogans was dependent on effective bacterial internalization and processing we hypothesized that a cytosolic pattern recognition receptor may be responsible for initiating this response. Additionally, the role of cytosolic DNA sensing in the innate immune response to L. interrogans remains unknown. To examine this, BMDMs from mice deficient in cGAS ( Cgas −/− ) and STING ( Sting1 gt/gt ) were challenged with L. interrogans and Ifna and Ifnb transcript induction was assessed by qPCR. BMDM from Cgas −/− and Sting1 gt/gt mice showed a marked defect in the transcription of Ifna and Ifnb at 4 hours post-infection with L. interrogans compared to BMDMs from WT mice ( Fig 4A, B ). Consistent with this, BMDMs from Cgas −/− and Sting1 gt/gt mice also had diminished IFNβ secretion, but not TNF secretion, into culture supernatants as measured by ELISA compared to WT macrophages ( Fig 4C, D ). ISGs downstream of IFN signaling were also diminished in BMDMs from Cgas −/− and Sting1 gt/gt mice compared to WT following infection with L. interrogans ( Fig 4E ). A similar reduction in IFNB transcript levels was observed in the human monocytic THP-1 cell line with CGAS and STING1 gene knockout ( Fig 4F ) indicating L. interrogans can activate the cGAS-STING pathway in both mouse and humans. Furthermore, leptospiral DNA directly transfected into BMDMs induced IFNβ production in a cGAS and STING dependent manner ( Fig 4G ). Download figure Open in new tab Fig 4. L. interrogans induced type I IFN is dependent on cGAS-STING. (A, B) Relative expression of Ifna and Ifnb transcripts in BMDMs from WT, Cgas -/- , and Sting1 gt/gt mice 4h after infection with L. interrogans Fiocruz L1-130 (MOI 100). (C, D) Quantification of IFNβ and TNFα by ELISA in culture supernatants of WT, Cgas -/- , and Sting1 gt/gt BMDMs infected with L. interrogans Fiocruz L1-130 (MOI 100) for 4, 6 and 8h. (E) Relative expression of ifit1, ifit3, ifi44 , and zbp1 transcript levels in BMDMs infected with L. interrogans (MOI 100) at 6h post-infection. (F) Relative expression of IFNA and IFNB transcript levels in PMA-differentiated human THP-1 cells infected with L. interrogans (MOI 100) at 4h post infection. (G) Quantification of IFNβ by ELISA in culture supernatants of WT, Cgas -/- , and Sting1 gt/gt BMDMs transfected with 500 ng of L. interrogans DNA using Xfect transfection agent. (A-G) Data are pooled from three experiments and expressed as the mean ± SEM. Statistical significance was calculated by two-way ANOVA; *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001, ns=non-significant. To confirm that the differences in type I IFN responses seen in the absence of cGAS and STING is not due to differences in phagocytosis pathways we visualized the number of bacteria either bound or internalized by BMDMs. We found similar numbers of bacteria bound to or internalized by WT, Cgas −/− and Sting1 gt/gt BMDMs at 3 hours post-infection ( S2 Fig ). Taken together these data demonstrate that L. interrogans activates the cGAS-STING signaling cascade thereby inducing secretion of type I IFN and the subsequent transcription of ISGs in macrophages. Furthermore, as the production of type I IFN is cGAS dependent, these data suggest that cytosolic recognition of dsDNA initiates this pathway as opposed to bacterial derived cyclic dinucleotides. STING is required for effective control of L. interrogans in mice in vivo To evaluate the contribution of STING in the control of L. interrogans in vivo , we intraperitoneally infected WT and Sting1 gt/gt mice with L. interrogans and assessed bacterial burdens in the spleen and kidneys 3 days post-infection ( Fig 5A ). Higher bacterial burdens were observed in the spleen of Sting1 gt/gt mice compared to WT ( Fig 5B ). No significant difference in bacterial burden was observed in the kidneys between WT and Sting1 gt/gt mice ( Fig 5C ). Lower expression of Ifna and Ifnb was seen in the spleen of Sting1 gt/gt mice compared to WT at 24 hours post-infection ( Fig 5D ). To assess renal colonization by L. interrogans , WT and Sting1 gt/gt mice were subcutaneously infected with 1×10 6 spirochetes and kidney colonization assessed at 14 days post-infection ( Fig 5E ). Sting1 gt/gt mice had higher kidney colonization with L. interrogans compared to WT mice ( Fig 5F ). Taken together these data suggest that cytosolic recognition of L. interrogans activates the cGAS-STING pathway to control L. interrogans replication in vivo and limits chronic kidney colonization. Download figure Open in new tab Fig 5. Increased susceptibility of STING-deficient mice to L. interrogans in vivo . (A) Schematic of acute infection model; WT and Sting1 gt/gt mice were infected intraperitoneally with 2×10 8 CFU of L. interrogans Fiocruz L1-130. (B, C) Bacterial abundance in the spleen and kidney at day 3 post-infection was assessed by qPCR for the L. interrogans lipL32 gene and normalized to the eukaryotic PPIA gene. Data are pooled from two independent experiments: n=10 WT, n=9 Sting1 gt/gt . (D) Relative expression of Ifna, Ifnb , and tnf transcripts in spleen 24h post-acute infection in WT and Sting1 gt/gt mice. n=5 WT, n=5 Sting1 gt/gt . Statistical significance was calculated by two-way ANOVA. (E) Schematic of chronic infection model; WT and Sting1 gt/gt mice were infected subcutaneously with 1×10 6 CFU of L. interrogans Fiocruz L1-130. (H) Bacterial abundance in the kidney at day 14 post-infection was assessed by qPCR as above. Data are pooled from two independent experiments: n=6 WT, n=6 Sting1 gt/gt . (B, C, F) Statistical significance calculated by Mann-Whitney U test. *p<0.05, **p<0.01, ns=non-significant. Discussion In this work, we have shown that L. interrogans induces a robust type I IFN response from human and mouse macrophages. The induction of type I IFN by L. interrogans was dependent on the cGAS-STING pathway as macrophages deficient in either cGAS or STING had markedly diminished expression of type I IFN and ISGs. Interestingly, the L. interrogans mediated production of type I IFN required bacterial phagocytosis and phagosomal acidification. We further demonstrate that the induction of type I IFN plays an important role in the host response to infection with L. interrogans in vivo ; mice deficient in STING or IFNAR displayed increased susceptibility to L. interrogans infection and had increased kidney colonization at 14 days after subcutaneous challenge. Leptospires are largely thought to be extracellular pathogens; however, studies have shown leptospires to be found within macrophages ( 19 , 21 ). The fate and impact of these intracellular leptospires has been unclear. Santecchia et al. demonstrated that L. interrogans can actively enter into both mouse and human macrophages ( 21 ). Surprisingly, they also found that the leptospires did not replicate within macrophages but could rapidly exit these cells intact ( 21 ). Using a pH-sensitive dye to label L. interrogans , we found that leptospires were being taken up in acidic sub-cellular compartments in macrophages. A fraction of internalized leptospires colocalized with Rab5 and LAMP1, suggesting the presence of leptospires in phagocytic vesicles. We further found that the pharmacologic inhibitors cytochalasin D, bafilomycin A and chymostatin, that target phagocytic uptake, phagosomal acidification, and proteolytic activity, respectively, inhibited L. interrogans induced IFNβ production. These data suggest that the phagocytosis and phagosomal processing of L. interrogans is required for the activation of the cGAS-STING pathway. It remains unclear to what extent these internalized leptospires are killed or damaged resulting in bacterial DNA release into the macrophage cytosol. The cGAS-STING pathway can drive the production of type I IFN in response to pathogen-derived DNA as well as endogenous host DNA ( 50 – 53 ). cGAS can bind to dsDNA and produce cGAMP, which then binds to and activates STING on the endoplasmic reticulum membrane ( 51 , 53 – 55 ). This results in TBK1 phosphorylation, which phosphorylates the transcription factor IRF3 and drives the induction of type I IFN ( 56 – 58 ). We observed that Leptospira induces a type I IFN response, coinciding with the phosphorylation of TBK1 and IRF3, supporting a role for cGAS-STING in the detection of intracellular L. interrogans . Several intracellular bacterial pathogens, including Salmonella enterica serovar Typhimurium, L. monocytogenes and M. tuberculosis have been shown to activate the cGAS-STING pathway to drive type I IFN production ( 23 , 25 , 59 ). Extracellular bacteria, including Klebsiella pneumoniae and Pseudomonas aeruginosa , have also been shown to activate the cGAS-STING pathway ( 26 ). The spirochete B. burgdorferi can activate the cGAS-STING pathway in macrophages and fibroblasts ( 29 ). Although cGAS-STING did not appear to play a role in control of B. burgdorferi burdens in vivo , inflammation and joint pathology were diminished in cGAS-deficient mice infected with B. burgdorferi ( 29 ). The mechanism by which cGAS-STING recognizes L. interrogans remains unclear. Bacterial derived cyclic dinucleotides have been shown to directly activate STING ( 30 – 32 ). Although L. interrogans can produce cyclic dinucleotides ( 34 , 35 ), our finding that cGAS is required for L. interrogans -driven type I IFN production suggests that sensing of cytosolic DNA is required to engage the cGAS-STING pathway in response to L. interrogans and that L. interrogans derived cyclic dinucleotides do not directly activate STING. As internalization and phagocytic processing of L. interrogans is required for type I IFN production by macrophages it is possible that internalized spirochetes either shed DNA or are damaged within the phagosome resulting in release of bacterial DNA that can be recognized by cytosolic cGAS. Another possibility is that infection of macrophages with L. interrogans results in mitochondrial stress resulting in the release of mtDNA which can then activate cGAS-STING ( 59 ). It should be noted that other pattern recognition receptors, including RIG-I/MDA5/MAVS, TLR3, TLR4, TLR7, TLR8, and TLR9, may be involved in the induction of type I IFN responses ( 60 – 67 ). The role of these pattern recognition receptors in driving the type I IFN response to L. interrogans will require further investigation. The murine model of leptospirosis remains poorly characterized, leaving many questions unanswered regarding the host response to Leptospira spp. A recent study by Papadopoulos et al. on the progression of acute leptospirosis in mouse models provides valuable insights into how severe leptospirosis in mice mimics aspects of human disease ( 68 ). This study attributes the observed mouse mortality in the acute leptospirosis model to severe myocarditis and neutrophil infiltration, in the absence of a significant cytokine storm ( 68 ). Our in vivo infection of Sting1 gt/gt and Ifnar1 -/- mice with L. interrogans revealed a role for type I IFN in both the early control of bacterial replication as well as in chronic kidney colonization with L. interrogans . The effect of the L. interrogans -driven type I IFN response on neutrophil function in vivo remains to be evaluated. In addition, the contribution of other cell types, such as renal tubular epithelial cells, in the production of type I IFN in response to L. interrogans will be important to determine. In conclusion, our study is the first to show that L. interrogans activates the cytosolic cGAS-STING DNA sensing pathway in mouse and human macrophages resulting in a type I IFN response. Future studies are required to determine the source of cytosolic DNA that is recognized by cGAS-STING following L. interrogans infection of macrophages. Additionally, determining the intracellular fate of L. interrogans will be important in understanding if bacterial damage and release of bacterial DNA is a prerequisite for activation of cGAS-STING. We also found that activation of STING and signaling through IFNAR are required for controlling L. interrogans replication and kidney colonization in mice in vivo. Evaluating the role of type I IFN production during the course of human disease remains an unresolved question and may provide insights into species specific responses. Materials and methods Ethics statement All studies with mice were performed in accordance with the recommendations in the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health and were reviewed and approved by the Institutional Animal Care committee at Cedars-Sinai Medical Center (CSMC IACUC #006777). Mice Wild-type (WT) C57BL/6J (JAX stock# 000664) mice were purchased from Jackson laboratories and used as controls unless otherwise stated. The generation of Sting1 gt/gt (JAX stock# 017537), Cgas -/- (JAX stock# 026554) and Ifnar1 -/- (JAX stock# 028288) have been previously described ( 69 – 71 ). Mice were bred and maintained in a specific-pathogen free facility. Both male and female mice 6–12 wk of age were used; however, mice were sex and age matched for individual experiments. Bacterial culture Leptospira interrogans serovar Copenhageni strain Fiocruz L1-130 and Leptospira biflexa serovar Patoc strain Patoc 1 were grown in Hornsby-Alt-Nally (HAN) media ( 72 ). Cultures were incubated at 30°C and passaged weekly. L. interrogans was passaged a maximum of five times after collection from kidneys of infected Golden Syrian hamsters. Culture densities were determined by diluting 10 μL of culture into phosphate-buffered saline (PBS) and counting by darkfield microscopy with a Zeiss Axio Lab A1 or AmScope B340 Series microscope. Prior to experiments, leptospires were centrifuged at 9,000 x g for 4 min at room temperature. For in vitro experiments, leptospires were resuspended in DMEM to a final density of 1 x 10 9 /mL. For heat killing, the leptospires were heated at 56°C for 30 min. For labelling of bacteria with pHrodo™ Red, succinimidyl ester (Invitrogen # P36600), leptospires were incubated for 45 min at room temperature with 0.1 mM dye. Labelled bacteria were washed and then incubated with BMDMs to analyze increase in fluorescence using an Incucyte SX5 Live-Cell Analysis System (Sartorius). For in vivo experiments, leptospires were resuspended in HAN media to a density of 1 x 10 9 /mL following centrifugation of the culture. After resuspension, 200 μL was used to inoculate mice for acute infection; for renal colonization, the cell suspension was diluted 100-fold prior to inoculation of mice with 100 μL. In vitro stimulation of bone marrow–derived macrophages Bone marrow–derived macrophages (BMDMs) were generated from cells collected by flushing mouse femurs and tibias and culturing the cells in 10 mm dish for 6-7 days in DMEM (Corning #10-013-CV) with 10% FBS (R&D systems #S12450H), penicillin-streptomycin (Gibco # 15140122), and L-glutamine (Gibco #25030-081) in the presence of 20% L929 cell-conditioned medium. Upon differentiation, BMDMs were harvested using Versene (Gibco# 15040066) and used for subsequent experiments. BMDMs were either left unstimulated or infected with L. interrogans Fiocruz L1-130 or L. biflexa Patoc I at multiplicity of infection (MOI) of 100 for 4, 6, and 8 h. Wherever indicated, BMDMs were pre-incubated for 45-60 mins with 2µM cytochalasin D (Sigma-Aldrich #C8273), 400nM bafilomycin A (Invivogen #tlrl-baf1) and 100µM chymostatin (Caymen Chemical #15114) before addition of bacteria. For transfection of leptospiral DNA, 500 ng of DNA isolated from L. interrogans Fiocruz L1-130 cultures was incubated with Xfect transfection reagent (Takara Bio #631317) for 10 min and BMDMs were treated with the mixture for 6 h. After respective stimulations, supernatants were collected for quantification of cytokines and cells were preserved in RNAlater Stabilization Solution (Invitrogen #AM7020) for RNA isolation using RNeasy Mini kit (Qiagen #74106) as per the manufacturers’ instruction. Cytokines and chemokines were quantified in the cell culture supernatants using mouse ELISA kit for TNF (Invitrogen #88-7324-88), IFNβ (R&D Systems #DY8234), IL-10 (R&D Systems #DY417) and IL-6 (R&D Systems #DY406) following the manufacturers’ protocol. Immunofluorescence staining and confocal microscopy 3×10 4 BMDMs were seeded in an 8-well Millicell EZ Slide (Millipore Sigma #PEZGS0816) and incubated for 3 h with L. interrogans Fiocruz L1-130 at MOI 100 or left unstimulated. Cells were washed with PBS three times and fixed with 4% PFA (Electron Microscopy Sciences #15710) for 10 minutes at room temperature. For analyzing bound bacteria, after fixation, cells were not permeabilized and proceeded with immunofluorescence (IF) staining. For analyzing total bound and internalized bacteria, fixed cells were permeabilized using 0.1% TritonX-100 (VWR Life Science # 9002-93-1) for 10 min. Cells were then incubated in IF blocking buffer containing 2.5% BSA (RPI # A30075) and normal donkey serum (Abcam #ab7475) in PBS (Corning #21-040-CV) for 1 h at room temperature. Primary antibodies: rat anti-mouse LAMP1 (clone 1D4B, eBioscience), mouse anti-mouse Rab5 (clone rab5-65, Sigma-Aldrich), and rabbit anti-sera raised against L. interrogans Fiocruz L1-130 were diluted in IF blocking buffer and added to cells for overnight incubation at 4°C. Cells were washed three times with PBS and AF488 anti-rabbit IgG, AF555 anti-mouse IgG, and AF647 anti-rat IgG diluted in IF buffer were incubated with the cells for 1 h at room temperature. Cells were washed three times with PBS before removing the well borders. VECTASHIELD Vibrance Antifade Mounting Medium with DAPI (Vector Laboratories #H-1800) was added on top of each well of stained cells and sealed with a coverslip. Imaging was done using either Zeiss Z780 confocal microscope or Leica Stellaris confocal microscope. For counting the number of bacterial puncta that are bound vs the total number of bacteria bound and internalized, the Image J manual cell counting tool was used. For analyzing the percentage of total bacteria that colocalizes with Rab5 or LAMP1, the JaCoP pixel-based colocalization tool was used to calculate the Manders’ coefficient using manual thresholding. In vivo infections For acute infection, mice were injected intraperitoneally with 2×10 8 bacteria and on day 3 following infection spleen and kidney were harvested and stored in RNAlater Stabilization Solution (Invitrogen #AM7020) for further processing. Tissues were weighed and homogenized in PBS with 2.8 mm ceramic beads (Omni #19-646) using a FastPrep-24 Classic bead beating grinder and lysis system (MP # 116004500). Subsequently DNA was extracted from appropriate tissue amount using DNeasy Blood and Tissue kit (Qiagen # 69506) using the manufacturers’ protocol. To assess kidney colonization, mice were injected subcutaneously with 1×10 6 bacteria and on day 14 following infection the kidney was harvested and processed as above. qPCR cDNA was synthesized from the isolated RNA using PrimeScript RT master mix (Takara Bio #RR036A) as per the manufacturers’ protocol. Transcripts were amplified with the primers listed in S1 Table using PowerUP SYBR Green PCR master mix (Applied Biosystems #25742) in Applied biosystems ViiA7 96 well system. The transcript levels were normalized to housekeeping gene β-actin and data are shown as fold change of transcripts relative to mean of WT group using the ΔΔ threshold cycle (Ct) method. For quantifying the relative abundance of bacteria in kidneys and spleen of infected mice, the DNA was amplified with primers specific to the lipL32 gene for L. interrogans and normalized to eukaryotic tissue inputs by amplification of PPIA housekeeping gene. Data are shown as relative abundance of bacteria normalized to tissue inputs and compared to the WT group on day 3 post infection using the ΔΔ Ct method. Cell lines Human monocyte cell line THP-1 with CGAS and STING1 gene knockout were generated by CRISPR/Cas9 as previously described ( 73 ). THP-1 monocytes were grown in RPMI-1640 medium (ATCC #30-2001) with 10% FBS (R&D systems #S12450H), penicillin-streptomycin (Gibco # 15140122), L-glutamine (Gibco #25030-081) and β-mercaptoethanol (Thermo Fisher # 21985023). THP-1 monocytes were differentiated using 50 nM PMA for 3-4 h, followed by an overnight rest period. The cells were then infected with L. interrogans Fiocruz L1-130 at MOI 100; 4 h later the cells were harvested for RNA isolation. Western blotting Cell lysates were prepared in radioimmunoprecipitation assay (RIPA) lysis buffer (Cell Signaling Technology #9806S) with Halt Protease and Phosphatase Inhibitor Cocktail (Thermo scientific #78440). Proteins were separated on NuPAGE Bis-Tris Mini Protein Gels, 4–12%, 1.0–1.5 mm and transferred to a polyvinylidene difluoride (PVDF) membrane using either the XCell II blotting system (Invitrogen #EI0002) or iBlot 2 Transfer system (Invitrogen #IB24002). Membranes were blocked with 1% BSA in tris-buffered saline (TBS) with 0.1% Tween-20 and incubated with primary antibody overnight at 4°C. Primary antibodies from Cell signaling technologies (CST) were used as follows: TBK1 (CST #3504S), p-TBK1 (CST #5483S), IRF3 (CST #4302S), p-IRF3 (CST #4947S) and β-actin (CST #4970S). After washing, membranes were incubated with HRP-tagged anti-rabbit IgG (Cell Signaling Technology #7074S). Membranes were developed using SuperSignal West Pico PLUS (Thermo Scientific #34580) or Femto substrate (Thermo Scientific #34094) and imaged in LI-COR Odyssey Fc. Statistics Data were graphed and the indicated statistical tests performed using GraphPad Prism 9 software. If not otherwise stated, analysis was performed with Unpaired t test with Welch’s correction for single comparison or Two-way ANOVA for multiple comparisons. Author contributions Conceptualization, S.G., S.L.C., F.S.S. and D.A.H.; Methodology, S.G., J.M., J.C., F.S.S. and D.A.H.; Investigation, S.G., J.M. and B.R.; Writing – Original Draft, S.G., F.S.S. S.L.C. and D.A.H.; Writing – Review & Editing, S.G., F.S.S. S.L.C. and D.A.H.; Funding Acquisition, S.L.C., J.D., F.S.S. and D.A.H.; Resources, A.P.W., N.K., C.S. and A.D.; Supervision, S.L.C., F.S.S. and D.A.H. Figure Legends Download figure Open in new tab S1 Fig. Analysis of IFNβ production from BMDM infected with Leptospira. (A) Dose dependent induction of IFNβ was quantified by ELISA in culture supernatants of WT BMDMs infected with L. interrogans (MOI 1, 10, 100) for 4, 6, and 8h. Data are pooled from two independent experiments and expressed as the mean ± SD. (B) Quantification of IFNβ levels by ELISA in culture supernatants of WT BMDM infected with live or heat-killed L. interrogans (MOI 100) at 6h post-infection. Data are pooled from three independent experiments. Statistical significance was calculated by unpaired Student’s t test. (C) Quantification of IFNβ levels by ELISA in culture supernatants of WT BMDM infected with L. interrogans Fiocruz L1-130 (MOI 100) or L. biflexa Patoc 1 (MOI 100) at 4, 6, and 8h post-infection. Statistical significance calculated by two-way ANOVA. ns=non-significant. Download figure Open in new tab S2 Fig. Analysis of L. interrogans binding and internalization by BMDM. Quantification of immunofluorescence images of bacteria bound or total bacteria (bound and internalized) in WT, Cgas -/- and Sting1 gt/gt BMDMs at 3h post infection with L. interrogans (MOI 100). Data are pooled from three frames per experiment from three independent experiments. Statistical significance was calculated by two-way ANOVA. ns=non-significant. View this table: View inline View popup Download powerpoint S1 Table: Primers used for qPCR Acknowledgments This work was supported by National Institutes of Health grants P01 AI168148 (D.A.H.), R01 AI175101 (F.S.S. and J.D.), and R01 AI177233 (F.S.S. and S.L.C.). References 1. ↵ Giraud-Gatineau A , Nieves C , Harrison LB , Benaroudj N , Veyrier FJ , Picardeau M . Evolutionary insights into the emergence of virulent Leptospira spirochetes . PLOS Pathog . 2024 Jul 17; 20 ( 7 ): e1012161 . OpenUrl CrossRef PubMed 2. ↵ Ellis WA . Animal leptospirosis . Curr Top Microbiol Immunol . 2015 ; 387 : 99 – 137 . OpenUrl CrossRef PubMed 3. ↵ Ko AI , Goarant C , Picardeau M . Leptospira: The dawn of the molecular genetics era for an emerging zoonotic pathogen . Nat Rev Microbiol . 2009 Oct ; 7 ( 10 ): 736 – 47 . OpenUrl CrossRef PubMed Web of Science 4. Ratet G , Veyrier FJ , d’Andon MF , Kammerscheit X , Nicola MA , Picardeau M , et al. Live Imaging of Bioluminescent Leptospira interrogans in Mice Reveals Renal Colonization as a Stealth Escape from the Blood Defenses and Antibiotics . PLoS Negl Trop Dis . 2014 Dec 4; 8 ( 12 ): e3359 . OpenUrl CrossRef PubMed 5. ↵ Haake DA , Levett PN . Leptospirosis in Humans . In: Adler B , editor. Leptospira and Leptospirosis [Internet] . Berlin, Heidelberg : Springer ; 2015 [cited 2025 Apr 15]. p. 65 – 97 . Available from : doi: 10.1007/978-3-662-45059-8_5 OpenUrl CrossRef PubMed 6. ↵ Costa F , Hagan JE , Calcagno J , Kane M , Torgerson P , Martinez-Silveira MS , et al. Global Morbidity and Mortality of Leptospirosis: A Systematic Review . PLoS Negl Trop Dis . 2015 Sep 17; 9 ( 9 ): e0003898 . OpenUrl CrossRef PubMed 7. ↵ Holzapfel M , Bonhomme D , Cagliero J , Vernel-Pauillac F , Fanton d’Andon M , Bortolussi S , et al. Escape of TLR5 Recognition by Leptospira spp.: A Rationale for Atypical Endoflagella . Front Immunol . 2020 ; 11 :2007. 8. ↵ Ratet G , Santecchia I , d’Andon MF , Vernel-Pauillac F , Wheeler R , Lenormand P , et al. LipL21 lipoprotein binding to peptidoglycan enables Leptospira interrogans to escape NOD1 and NOD2 recognition . PLOS Pathog . 2017 Dec 6; 13 ( 12 ): e1006725 . OpenUrl CrossRef PubMed 9. ↵ Werts C , Tapping RI , Mathison JC , Chuang TH , Kravchenko V , Saint Girons I , et al. Leptospiral lipopolysaccharide activates cells through a TLR2-dependent mechanism . Nat Immunol . 2001 Apr ; 2 ( 4 ): 346 – 52 . OpenUrl CrossRef PubMed Web of Science 10. ↵ Hsu SH , Hung CC , Chang MY , Ko YC , Yang HY , Hsu HH , et al. Active Components of Leptospira Outer Membrane Protein LipL32 to Toll-Like Receptor 2 . Sci Rep . 2017 Aug 21; 7 ( 1 ): 8363 . OpenUrl CrossRef PubMed 11. ↵ Murray GL , Srikram A , Henry R , Hartskeerl RA , Sermswan RW , Adler B . Mutations affecting Leptospira interrogans lipopolysaccharide attenuate virulence . Mol Microbiol . 2010 Nov ; 78 ( 3 ): 701 – 9 . OpenUrl CrossRef PubMed Web of Science 12. ↵ Nahori MA , Fournié-Amazouz E , Que-Gewirth NS , Balloy V , Chignard M , Raetz CRH , et al. Differential TLR recognition of leptospiral lipid A and lipopolysaccharide in murine and human cells . J Immunol Baltim Md 1950 . 2005 Nov 1; 175 ( 9 ): 6022 – 31 . OpenUrl 13. ↵ Chassin C , Picardeau M , Goujon JM , Bourhy P , Quellard N , Darche S , et al. TLR4- and TLR2-mediated B cell responses control the clearance of the bacterial pathogen , Leptospira interrogans. J Immunol Baltim Md 1950 . 2009 Aug 15; 183 ( 4 ): 2669 – 77 . OpenUrl 14. ↵ Tu V , Adler B , Faine S . The role of macrophages in the protection of mice against leptospirosis: in vitro and in vivo studies . Pathology (Phila ). 1982 Oct ; 14 ( 4 ): 463 – 8 . OpenUrl 15. Isogai E , Kitagawa H , Isogai H , Kurebayashi Y , Ito N . Phagocytosis as a defense mechanism against infection with leptospiras . Zentralblatt Für Bakteriol Mikrobiol Hyg Ser Med Microbiol Infect Dis Virol Parasitol . 1986 Feb 1; 261 ( 1 ): 65 – 74 . OpenUrl 16. ↵ Santecchia I , Vernel-Pauillac F , Rasid O , Quintin J , Gomes-Solecki M , Boneca IG , et al. Innate immune memory through TLR2 and NOD2 contributes to the control of Leptospira interrogans infection . PLOS Pathog . 2019 May 20; 15 ( 5 ): e1007811 . OpenUrl CrossRef PubMed 17. ↵ Ferrer MF , Scharrig E , Charo N , Rípodas AL , Drut R , Carrera Silva EA , et al. Macrophages and Galectin 3 Control Bacterial Burden in Acute and Subacute Murine Leptospirosis That Determines Chronic Kidney Fibrosis . Front Cell Infect Microbiol [Internet] . 2018 Oct 30 [cited 2024 Oct 24]; 8 . Available from: https://www.frontiersin.org/journals/cellular-and-infection-microbiology/articles/10.3389/fcimb.2018.00384/full 18. ↵ Merien F , Baranton G , Perolat P . Invasion of Vero cells and induction of apoptosis in macrophages by pathogenic Leptospira interrogans are correlated with virulence . Infect Immun . 1997 Feb ; 65 ( 2 ): 729 – 38 . OpenUrl Abstract / FREE Full Text 19. ↵ Toma C , Okura N , Takayama C , Suzuki T . Characteristic features of intracellular pathogenic Leptospira in infected murine macrophages . Cell Microbiol . 2011 ; 13 ( 11 ): 1783 – 92 . OpenUrl CrossRef PubMed 20. ↵ Davis JM , Haake DA , Ramakrishnan L . Leptospira interrogans Stably Infects Zebrafish Embryos, Altering Phagocyte Behavior and Homing to Specific Tissues . PLoS Negl Trop Dis . 2009 Jun 23; 3 ( 6 ): e463 . OpenUrl CrossRef PubMed 21. ↵ Santecchia I , Bonhomme D , Papadopoulos S , Escoll P , Giraud-Gatineau A , Moya-Nilges M , et al. Alive Pathogenic and Saprophytic Leptospires Enter and Exit Human and Mouse Macrophages With No Intracellular Replication . Front Cell Infect Microbiol . 2022 ; 12 : 936931 . OpenUrl CrossRef PubMed 22. ↵ Cai X , Chiu YH , Chen ZJ . The cGAS-cGAMP-STING Pathway of Cytosolic DNA Sensing and Signaling . Mol Cell . 2014 Apr 24; 54 ( 2 ): 289 – 96 . OpenUrl CrossRef PubMed Web of Science 23. ↵ Watson RO , Bell SL , MacDuff DA , Kimmey JM , Diner EJ , Olivas J , et al. The Cytosolic Sensor cGAS Detects Mycobacterium tuberculosis DNA to Induce Type I Interferons and Activate Autophagy . Cell Host Microbe . 2015 Jun 10; 17 ( 6 ): 811 – 9 . OpenUrl CrossRef PubMed 24. Gao D , Wu J , Wu YT , Du F , Aroh C , Yan N , et al. Cyclic GMP-AMP Synthase Is an Innate Immune Sensor of HIV and Other Retroviruses . Science . 2013 Aug 23; 341 ( 6148 ): 903 – 6 . OpenUrl Abstract / FREE Full Text 25. ↵ Hansen K , Prabakaran T , Laustsen A , Jørgensen SE , Rahbæk SH , Jensen SB , et al. Listeria monocytogenes induces IFNβ expression through an IFI16-, cGAS- and STING-dependent pathway . EMBO J . 2014 Aug ; 33 ( 15 ): 1654 – 66 . OpenUrl Abstract / FREE Full Text 26. ↵ Zhou CM , Wang B , Wu Q , Lin P , Qin SG , Pu QQ , et al. Identification of cGAS as an innate immune sensor of extracellular bacterium Pseudomonas aeruginosa . iScience . 2021 Jan 22; 24 ( 1 ): 101928 . OpenUrl CrossRef PubMed 27. ↵ Patrick KL , Bell SL , Watson RO . For Better or Worse: Cytosolic DNA Sensing during Intracellular Bacterial Infection Induces Potent Innate Immune Responses . J Mol Biol . 2016 Aug 28; 428 ( 17 ): 3372 – 86 . OpenUrl CrossRef PubMed 28. ↵ Platanias LC . Mechanisms of type-I- and type-II-interferon-mediated signalling . Nat Rev Immunol . 2005 May ; 5 ( 5 ): 375 – 86 . OpenUrl CrossRef PubMed Web of Science 29. ↵ Farris LC , Torres-Odio S , Adams LG , West AP , Hyde JA . Borrelia burgdorferi Engages Mammalian Type I IFN Responses via the cGAS-STING Pathway . J Immunol Baltim Md 1950 . 2023 Jun 1; 210 ( 11 ): 1761 – 70 . OpenUrl 30. ↵ Chauvin SD , Stinson WA , Platt DJ , Poddar S , Miner JJ . Regulation of cGAS and STING signaling during inflammation and infection . J Biol Chem . 2023 Jul 1; 299 ( 7 ): 104866 . OpenUrl CrossRef PubMed 31. Whiteley AT , Eaglesham JB , de Oliveira Mann CC , Morehouse BR , Lowey B , Nieminen EA , et al. Bacterial cGAS-like enzymes synthesize diverse nucleotide signals . Nature . 2019 Mar ; 567 ( 7747 ): 194 – 9 . OpenUrl CrossRef PubMed 32. ↵ Cohen D , Melamed S , Millman A , Shulman G , Oppenheimer-Shaanan Y , Kacen A , et al. Cyclic GMP–AMP signalling protects bacteria against viral infection . Nature . 2019 Oct ; 574 ( 7780 ): 691 – 5 . OpenUrl CrossRef PubMed 33. ↵ Liu H , Moura-Alves P , Pei G , Mollenkopf H , Hurwitz R , Wu X , et al. cGAS facilitates sensing of extracellular cyclic dinucleotides to activate innate immunity . EMBO Rep . 2019 Apr ; 20 ( 4 ): e46293 . OpenUrl Abstract / FREE Full Text 34. ↵ Thibeaux R , Soupé-Gilbert ME , Kainiu M , Girault D , Bierque E , Fernandes J , et al. The zoonotic pathogen Leptospira interrogans mitigates environmental stress through cyclic-di-GMP-controlled biofilm production . NPJ Biofilms Microbiomes . 2020 Jun 12; 6 : 24 . OpenUrl CrossRef PubMed 35. ↵ da Costa Vasconcelos FN , Maciel NK , Favaro DC , de Oliveira LC , Barbosa AS , Salinas RK , et al. Structural and Enzymatic Characterization of a cAMP-Dependent Diguanylate Cyclase from Pathogenic Leptospira Species . J Mol Biol . 2017 Jul 21; 429 ( 15 ): 2337 – 52 . OpenUrl CrossRef PubMed 36. ↵ Lacroix-Lamandé S , d’Andon M , Michel E , Ratet G , Philpott D , Girardin S , et al. Downregulation of the Na/K-ATPase Pump by Leptospiral Glycolipoprotein Activates the NLRP3 Inflammasome . J Immunol Baltim Md 1950 . 2012 Mar 15; 188 : 2805 – 14 . OpenUrl 37. ↵ Liu S , Cai X , Wu J , Cong Q , Chen X , Li T , et al. Phosphorylation of innate immune adaptor proteins MAVS, STING, and TRIF induces IRF3 activation . Science . 2015 Mar 13; 347 ( 6227 ): aaa2630 . OpenUrl Abstract / FREE Full Text 38. ↵ Henry RM , Hoppe AD , Joshi N , Swanson JA . The uniformity of phagosome maturation in macrophages . J Cell Biol . 2004 Jan 12; 164 ( 2 ): 185 – 94 . OpenUrl Abstract / FREE Full Text 39. Bucci C , Parton RG , Mather IH , Stunnenberg H , Simons K , Hoflack B , et al. The small GTPase rab5 functions as a regulatory factor in the early endocytic pathway . Cell . 1992 Sep 4; 70 ( 5 ): 715 – 28 . OpenUrl CrossRef PubMed Web of Science 40. Zimmerli S , Majeed M , Gustavsson M , Stendahl O , Sanan DA , Ernst JD . Phagosome-lysosome fusion is a calcium-independent event in macrophages . J Cell Biol . 1996 Jan ; 132 ( 1–2 ): 49 – 61 . OpenUrl Abstract / FREE Full Text 41. ↵ Duclos S , Diez R , Garin J , Papadopoulou B , Descoteaux A , Stenmark H , et al. Rab5 regulates the kiss and run fusion between phagosomes and endosomes and the acquisition of phagosome leishmanicidal properties in RAW 264.7 macrophages . J Cell Sci . 2000 Oct ; 113 Pt 19 : 3531 – 41 . OpenUrl Abstract / FREE Full Text 42. ↵ Bowman EJ , Siebers A , Altendorf K . Bafilomycins: a class of inhibitors of membrane ATPases from microorganisms, animal cells, and plant cells . Proc Natl Acad Sci . 1988 Nov ; 85 ( 21 ): 7972 – 6 . OpenUrl Abstract / FREE Full Text 43. Werner G , Hagenmaier H , Drautz H , Baumgartner A , Zähner H . METABOLIC PRODUCTS OF MICROORGANISMS . 224 BAFILOMYCINS, A NEW GROUP OF MACROLIDE ANTIBIOTICS PRODUCTION, ISOLATION, CHEMICAL STRUCTURE AND BIOLOGICAL ACTIVITY . J Antibiot (Tokyo) . 1984 ; 37 ( 2 ): 110 – 7 . OpenUrl CrossRef PubMed 44. ↵ Yoshimori T , Yamamoto A , Moriyama Y , Futai M , Tashiro Y . Bafilomycin A1, a specific inhibitor of vacuolar-type H(+)-ATPase, inhibits acidification and protein degradation in lysosomes of cultured cells . J Biol Chem . 1991 Sep 15; 266 ( 26 ): 17707 – 12 . OpenUrl Abstract / FREE Full Text 45. ↵ Okunishi H , Miyazaki M , Toda N . Evidence for a Putatively New Angiotensin II-generating Enzyme in the Vascular Wall . J Hypertens . 1984 Jun ; 2 ( 3 ): 277 . OpenUrl PubMed 46. ↵ Miyazaki M , Takai S , Jin D , Muramatsu M . Pathological roles of angiotensin II produced by mast cell chymase and the effects of chymase inhibition in animal models . Pharmacol Ther . 2006 Dec 1; 112 ( 3 ): 668 – 76 . OpenUrl CrossRef PubMed Web of Science 47. ↵ Hamond C , LeCount K , Browne AS , Anderson T , Stuber T , Hicks J , et al. Concurrent colonization of rodent kidneys with multiple species and serogroups of pathogenic Leptospira . Appl Environ Microbiol . 2023 Oct 31; 89 ( 10 ): e0120423 . OpenUrl CrossRef PubMed 48. Putz EJ , Nally JE . Investigating the Immunological and Biological Equilibrium of Reservoir Hosts and Pathogenic Leptospira: Balancing the Solution to an Acute Problem? Front Microbiol [Internet] . 2020 Aug 14 [cited 2025 Apr 15]; 11 . Available from: https://www.frontiersin.orghttps://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2020.02005/full 49. ↵ Picardeau M . Virulence of the zoonotic agent of leptospirosis: still terra incognita? Nat Rev Microbiol . 2017 May ; 15 ( 5 ): 297 – 307 . OpenUrl CrossRef PubMed 50. ↵ Chen Q , Sun L , Chen ZJ . Regulation and function of the cGAS–STING pathway of cytosolic DNA sensing . Nat Immunol . 2016 Oct ; 17 ( 10 ): 1142 – 9 . OpenUrl CrossRef PubMed 51. ↵ Zhang X , Wu J , Du F , Xu H , Sun L , Chen Z , et al. The Cytosolic DNA Sensor cGAS Forms an Oligomeric Complex with DNA and Undergoes Switch-like Conformational Changes in the Activation Loop . Cell Rep . 2014 Feb 13; 6 ( 3 ): 421 – 30 . OpenUrl CrossRef PubMed Web of Science 52. Li X , Shu C , Yi G , Chaton CT , Shelton CL , Diao J , et al. Cyclic GMP-AMP Synthase Is Activated by Double-Stranded DNA-Induced Oligomerization . Immunity . 2013 Dec 12; 39 ( 6 ): 1019 – 31 . OpenUrl CrossRef PubMed Web of Science 53. ↵ Wu J , Sun L , Chen X , Du F , Shi H , Chen C , et al. Cyclic GMP-AMP is an endogenous second messenger in innate immune signaling by cytosolic DNA . Science . 2013 Feb 15; 339 ( 6121 ): 826 – 30 . OpenUrl Abstract / FREE Full Text 54. Ishikawa H , Barber GN . STING is an endoplasmic reticulum adaptor that facilitates innate immune signalling . Nature . 2008 Oct ; 455 ( 7213 ): 674 – 8 . OpenUrl CrossRef PubMed Web of Science 55. ↵ Zhong B , Yang Y , Li S , Wang YY , Li Y , Diao F , et al. The Adaptor Protein MITA Links Virus-Sensing Receptors to IRF3 Transcription Factor Activation . Immunity . 2008 Oct 17; 29 ( 4 ): 538 – 50 . OpenUrl CrossRef PubMed Web of Science 56. ↵ Fitzgerald KA , McWhirter SM , Faia KL , Rowe DC , Latz E , Golenbock DT , et al. IKKε and TBK1 are essential components of the IRF3 signaling pathway . Nat Immunol . 2003 May ; 4 ( 5 ): 491 – 6 . OpenUrl CrossRef PubMed Web of Science 57. Tanaka Y , Chen ZJ . STING Specifies IRF3 Phosphorylation by TBK1 in the Cytosolic DNA Signaling Pathway . Sci Signal . 2012 Mar 6; 5 ( 214 ): ra20 – ra20 . OpenUrl Abstract / FREE Full Text 58. ↵ Sharma S , tenOever BR , Grandvaux N , Zhou GP , Lin R , Hiscott J. Triggering the Interferon Antiviral Response Through an IKK-Related Pathway . Science . 2003 May 16; 300 ( 5622 ): 1148 – 51 . OpenUrl Abstract / FREE Full Text 59. ↵ Xu L , Li M , Yang Y , Zhang C , Xie Z , Tang J , et al. Salmonella Induces the cGAS-STING-Dependent Type I Interferon Response in Murine Macrophages by Triggering mtDNA Release . mBio . 2022 May 23; 13 ( 3 ): e03632 – 21 . OpenUrl PubMed 60. ↵ Goubau D , Deddouche S , Reis e Sousa C. Cytosolic Sensing of Viruses . Immunity . 2013 May 23; 38 ( 5 ): 855 – 69 . OpenUrl CrossRef PubMed Web of Science 61. Seth RB , Sun L , Ea CK , Chen ZJ . Identification and Characterization of MAVS, a Mitochondrial Antiviral Signaling Protein that Activates NF-κB and IRF3 . Cell . 2005 Sep 9; 122 ( 5 ): 669 – 82 . OpenUrl CrossRef PubMed Web of Science 62. Hoshino K , Sugiyama T , Matsumoto M , Tanaka T , Saito M , Hemmi H , et al. IκB kinase-α is critical for interferon-α production induced by Toll-like receptors 7 and 9 . Nature . 2006 Apr ; 440 ( 7086 ): 949 – 53 . OpenUrl CrossRef PubMed Web of Science 63. Uematsu S , Sato S , Yamamoto M , Hirotani T , Kato H , Takeshita F , et al. Interleukin-1 receptor-associated kinase-1 plays an essential role for Toll-like receptor (TLR)7- and TLR9-mediated interferon-α induction . J Exp Med . 2005 Mar 14; 201 ( 6 ): 915 – 23 . OpenUrl Abstract / FREE Full Text 64. Perry AK , Chow EK , Goodnough JB , Yeh WC , Cheng G . Differential Requirement for TANK-binding Kinase-1 in Type I Interferon Responses to Toll-like Receptor Activation and Viral Infection . J Exp Med . 2004 Jun 21; 199 ( 12 ): 1651 – 8 . OpenUrl Abstract / FREE Full Text 65. Hemmi H , Takeuchi O , Sato S , Yamamoto M , Kaisho T , Sanjo H , et al. The Roles of Two IκB Kinase-related Kinases in Lipopolysaccharide and Double Stranded RNA Signaling and Viral Infection . J Exp Med . 2004 Jun 21; 199 ( 12 ): 1641 – 50 . OpenUrl Abstract / FREE Full Text 66. Kawai T , Takahashi K , Sato S , Coban C , Kumar H , Kato H , et al. IPS-1, an adaptor triggering RIG-I- and Mda5-mediated type I interferon induction . Nat Immunol . 2005 Oct ; 6 ( 10 ): 981 – 8 . OpenUrl CrossRef PubMed Web of Science 67. ↵ Honda K , Taniguchi T . IRFs: master regulators of signalling by Toll-like receptors and cytosolic pattern-recognition receptors . Nat Rev Immunol . 2006 Sep ; 6 ( 9 ): 644 – 58 . OpenUrl CrossRef PubMed Web of Science 68. ↵ Papadopoulos S , Hardy D , Vernel-Pauillac F , Tichit M , Boneca IG , Werts C . Myocarditis and neutrophil-mediated vascular leakage but not cytokine storm associated with fatal murine leptospirosis . eBioMedicine . 2025 Feb 1; 112 : 105571 . OpenUrl CrossRef PubMed 69. ↵ Sauer JD , Sotelo-Troha K , von Moltke J , Monroe KM , Rae CS , Brubaker SW , et al. The N-ethyl-N-nitrosourea-induced Goldenticket mouse mutant reveals an essential function of Sting in the in vivo interferon response to Listeria monocytogenes and cyclic dinucleotides . Infect Immun . 2011 Feb ; 79 ( 2 ): 688 – 94 . OpenUrl Abstract / FREE Full Text 70. Schoggins JW , MacDuff DA , Imanaka N , Gainey MD , Shrestha B , Eitson JL , et al. Pan-viral specificity of IFN-induced genes reveals new roles for cGAS in innate immunity . Nature . 2014 Jan 30; 505 ( 7485 ): 691 – 5 . OpenUrl CrossRef PubMed Web of Science 71. ↵ Prigge JR , Hoyt TR , Dobrinen E , Capecchi MR , Schmidt EE , Meissner N . Type I IFNs act upon hematopoietic progenitors to protect and maintain hematopoiesis during Pneumocystis lung infection in mice . J Immunol Baltim Md 1950 . 2015 Dec 1; 195 ( 11 ): 5347 – 57 . OpenUrl 72. ↵ Hornsby RL , Alt DP , Nally JE . Isolation and propagation of leptospires at 37 °C directly from the mammalian host . Sci Rep . 2020 Jun 15; 10 ( 1 ): 9620 . OpenUrl CrossRef PubMed 73. ↵ Banerjee D , Langberg K , Abbas S , Odermatt E , Yerramothu P , Volaric M , et al. A non-canonical, interferon-independent signaling activity of cGAMP triggers DNA damage response signaling . Nat Commun . 2021 Oct 27; 12 ( 1 ): 6207 . OpenUrl CrossRef PubMed View the discussion thread. Back to top Previous Next Posted June 04, 2025. Download PDF Email Thank you for your interest in spreading the word about bioRxiv. NOTE: Your email address is requested solely to identify you as the sender of this article. Your Email * Your Name * Send To * Enter multiple addresses on separate lines or separate them with commas. You are going to email the following cGAS-STING dependent type I IFN protects against Leptospira interrogans renal colonization in mice Message Subject (Your Name) has forwarded a page to you from bioRxiv Message Body (Your Name) thought you would like to see this page from the bioRxiv website. Your Personal Message CAPTCHA This question is for testing whether or not you are a human visitor and to prevent automated spam submissions. Share cGAS-STING dependent type I IFN protects against Leptospira interrogans renal colonization in mice Suman Gupta , James Matsunaga , Bridget Ratitong , Sana Ismaeel , Diogo G. Valadares , A. Phillip West , Nagaraj Kerur , Christian Stehlik , Andrea Dorfleutner , Jargalsaikhan Dagvadorj , Jenifer Coburn , Andrea J. Wolf , Suzanne L. Cassel , David A. Haake , Fayyaz S. Sutterwala bioRxiv 2025.06.02.657349; doi: https://doi.org/10.1101/2025.06.02.657349 Share This Article: Copy Citation Tools cGAS-STING dependent type I IFN protects against Leptospira interrogans renal colonization in mice Suman Gupta , James Matsunaga , Bridget Ratitong , Sana Ismaeel , Diogo G. Valadares , A. Phillip West , Nagaraj Kerur , Christian Stehlik , Andrea Dorfleutner , Jargalsaikhan Dagvadorj , Jenifer Coburn , Andrea J. Wolf , Suzanne L. Cassel , David A. Haake , Fayyaz S. Sutterwala bioRxiv 2025.06.02.657349; doi: https://doi.org/10.1101/2025.06.02.657349 Citation Manager Formats BibTeX Bookends EasyBib EndNote (tagged) EndNote 8 (xml) Medlars Mendeley Papers RefWorks Tagged Ref Manager RIS Zotero Tweet Widget Facebook Like Google Plus One Subject Area Immunology Subject Areas All Articles Animal Behavior and Cognition (7633) Biochemistry (17681) Bioengineering (13890) Bioinformatics (41929) Biophysics (21446) Cancer Biology (18586) Cell Biology (25492) Clinical Trials (138) Developmental Biology (13374) Ecology (19897) Epidemiology (2067) Evolutionary Biology (24308) Genetics (15606) Genomics (22497) Immunology (17736) Microbiology (40385) Molecular Biology (17175) Neuroscience (88584) Paleontology (666) Pathology (2831) Pharmacology and Toxicology (4822) Physiology (7641) Plant Biology (15149) Scientific Communication and Education (2045) Synthetic Biology (4293) Systems Biology (9822) Zoology (2271)