Role of snx27 in protein trasnport and lipid signaling in cell models of polarized trafficking

Resumen

Sorting nexin 27 (SNX27) belongs to the SNX family of proteins that control intracellular protein trafficking. SNX27 bears a PX (Phox homology) domain that regulates its endosomal localization, a unique PDZ (PSD-95, Dlg1, ZO-1) domain and an atypical FERM (4.1, ezrin, radixin, moesin) domain, which bind short peptide sequence motifs in the cargo protein cytoplasmic domains. SNX27 recycles PDZ-interacting receptors at the neuronal synapse, and its low expression is associated with impaired synaptic function and neurological diseases. T lymphocyte antigen recognition leads to the formation of a highly organized structure termed immune synapse (IS), by analogy with the nervous synapse. SNX27-positive endosomes polarize to the IS, but the role of SNX27 during T cell activation remains unknown. In this study we investigated SNX27 dynamics and functions during IS formation. We characterized a previously unidentified FERM domain lipid-binding site that enhances SNX27 endosomal localization, and showed that a fraction of SNX27 accumulates at the IS in a PDZ ligand-dependent manner. We used proteomics to identify the SNX27 interactome in IS-forming T cells; a comparative interactome analysis of SNX27 WT and a mutant deficient for PDZ ligand recognition identified various cargoes linked to signaling and cytoskeletal regulation. We demonstrated that SNX27 controls the signaling and trafficking of some of these cargoes including the lipid kinase diacylglycerol kinase ζ (DGKζ) and the epithelial cell-cell junction protein zona occludens-2 (ZO-2). Proteomic analyses also detected the retromer and the actin nucleator WASH complex, and we confirmed that SNX27 acts as a WASH adaptor for PDZ cargoes in T cells. Analysis of Snx27−/− mice supported our studies in human T cell lines, underscoring a putative role for SNX27 in sustaining the function of the metabolic regulator mTOR (mammalian target of rapamycin) in activated T cells. This study broadens our knowledge of the SNX27 function in integrating sorting events with endosomal lipid signaling in T lymphocytes, and suggests conservation in the pathways that delimit polarized structures in nervous and epithelial systems during IS formation. For their migration and invasion, cancer cells also need to form highly organized structures. Cells assemble invadopodia to coordinate the signaling and trafficking that promote increased secretion and matrix degradation. The SNX27 partner WASH, assisted by the exocyst complex, controls integrin and metalloproteinase recycling at invadopodia. The role of SNX27 in invasive cell polarized recycling has nonetheless not been explored. We showed SNX27 localization to invadopodia in a cancer cell line, and addressed the effects of its downmodulation on invasive migration. We identified the phosphatidylinositol (4,5)-bisphosphate-synthesizing enzyme phosphatidylinositol 4-phosphate 5-kinase γ (PIP5K γ), a known modulator of exocyst function, as a SNX27 partner. Our results support a SNX27 function as an endosomal hub that assists lipid modulation in the recycling compartment, and suggest that the frequent amplification of SNX27 in human cancers promotes oncogenic traits in tumor cells.