Lipid Transfer Proteins and PI4KIIα Generate a Phosphoinositide-Linked Proteome

Summary Phosphoinositide (PIPn) lipid second messengers in membranes regulate a myriad of cellular processes. In the cytosol, the phosphatidylinositol (PI) 3-kinase (PI3K)/Akt pathway is scaffolded on IQGAP1 to facilitate the activation of Akt by the synthesis of PI3,4,5P3. In the nucleus, PIPn signaling occurs in regions devoid of membranes via their stable association with proteins. While several of these proteins have been identified, understanding the extent and impact of protein-linked PIPn signaling warrants further investigation. The tumor suppressor p53, was shown in the companion paper to be regulated by PI transfer proteins (PITPs) and a PI 4-kinase (PI4KIIα), which are required to form p53-PIPn complexes that assemble a nuclear PI3K/Akt pathway. Here we report that class I PITPs (PITPα/β) and PI 4-kinase initiate PIPn linkages to many different proteins. PITPα/β and PI4KIIα accumulate in the nucleoplasm in response to stress and are necessary to synthesize nuclear PIPns linked to proteins. These PITPα/β-dependent protein-PIPn complexes are detected by metabolically labeling cells with the PIPn precursor [H3]-myo-inositol and resist denaturation and SDS-PAGE, indicating that these protein-PIPn complexes represent a putative posttranslational modification. Proteomic analyses of proteins that are regulated by PITPα/β and/or are linked to PI4,5P2 have identified an emerging PIPylome that is enriched in metabolic, signaling, cytoskeletal and DNA repair pathway components. Taken together, these data provide evidence for an emerging proteome with linked PIPns that represent a PIPn signaling paradigm that is distinct from the membrane-localized pathway but utilizes many of the same PIP kinases and phosphatases. In brief Phosphatidylinositol transfer proteins and PI 4-kinase initiate a PIPn-linked protein network in membrane-free regions. Competing Interest Statement The authors have declared no competing interest. Footnotes ↵6 Co-first authors