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Macroautophagy, the degradation and recycling of cytosolic components in the lysosome, is an important cellular mechanism. It is a membrane-mediated process that is linked to vesicular trafficking events. The sorting nexin (SNX) protein family controls the sorting of a large array of cargoes, and various SNXs impact autophagy. To improve our understanding of their functions in vivo, we screened all Drosophila SNXs using inducible RNA interference in the fat body. Significantly, depletion of snazarus (snz) led to decreased autophagic flux. Interestingly, we observed altered distribution of Vamp7-positive vesicles with snz depletion, and snz's roles were conserved in human cells. SNX25, the closest human ortholog to snz, regulates both VAMP8 endocytosis and lipid metabolism. Through knockout-rescue experiments, we demonstrate that these activities are dependent on specific SNX25 domains and that the autophagic defects upon SNX25 loss can be rescued by ethanolamine addition. We also demonstrate the presence of differentially spliced forms of SNX14 and SNX25 in cancer cells. This work identifies a conserved role for snz/SNX25 as regulators of autophagic flux and reveals differential isoform expression between paralogs.

27Adipocytes store nutrients as lipid droplets (LDs), but how they organize their LD stores 28 to balance lipid uptake, storage, and mobilization remains poorly understood. Here, 29using Drosophila fat body (FB) adipocytes we characterize spatially distinct LD 30 populations that are maintained by different lipid pools. We identify peripheral LDs 31 (pLDs) that make close contact with the plasma membrane (PM) and are maintained by 32 lipophorin-dependent lipid trafficking. pLDs are distinct from larger cytoplasmic medial 33 LDs (mLDs) which are maintained by FASN1-dependent de novo lipogenesis. We find 34 that sorting nexin CG1514/Snazarus (Snz) associates with pLDs and regulates LD 35 homeostasis at ER-PM contact sites. Loss of SNZ perturbs pLD organization whereas 36Snz over-expression drives LD expansion, triacylglyceride production, starvation 37 resistance, and lifespan extension through a DESAT1-dependent pathway. We propose 38that Drosophila adipocytes maintain spatially distinct LD populations and identify Snz as 39 a novel regulator of LD organization and inter-organelle crosstalk. 40 41

Autophagy, the degradation and recycling of cytosolic components in the lysosome, is an essential cellular mechanism. It is a membrane-mediated process that is linked to vesicular trafficking events. The sorting nexin (SNX) protein family controls the sorting of a large array of cargoes, and various SNXs can impact autophagy. To gain a better understanding of their functions in vivo under nutrient starvation, we screened all Drosophila SNXs by RNAi in the fat body. Significantly, depletion of snazarus (snz) strongly impacted autolysosome formation and led to decreased autophagic flux. Interestingly, we observed altered distribution of Vamp7-positive vesicles with snz depletion and snz roles were conserved in human cells. SNX25 is the closest ortholog to snz, and we demonstrate a role for it in VAMP8 trafficking. We found that this activity was dependent on the SNX25 PX domain, and independent of SNX25 anchoring at the ER. We also demonstrate that differentially spliced forms of SNX14 and SNX25 are present in cancer cells. This work identifies a conserved role for snz/SNX25 as regulators of autophagic flux, and show differential isoform expression between orthologs.

No abstract