%0 Journal Article %1 BeerE1127 %A Beer, Katharina B. %A Rivas-Castillo, Jennifer %A Kuhn, Kenneth %A Fazeli, Gholamreza %A Karmann, Birgit %A Nance, Jeremy F. %A Stigloher, Christian %A Wehman, Ann M. %D 2018 %I National Academy of Sciences %J Proceedings of the National Academy of Sciences %K myown %N 6 %P E1127--E1136 %R 10.1073/pnas.1714085115 %T Extracellular vesicle budding is inhibited by redundant regulators of TAT-5 flippase localization and phospholipid asymmetry %U https://www.pnas.org/content/115/6/E1127 %V 115 %X Cells must interact with their environment to survive. The lipids and proteins of the plasma membrane send and receive signals at the cell surface to respond to stimuli. When the lipid bilayer of the plasma membrane is damaged, cells release membrane-bound extracellular vesicles to repair the membrane. Cells also release signals on extracellular vesicles to communicate at a distance. Here, we identify proteins that regulate the formation of extracellular vesicles from the plasma membrane, providing additional tools to control their release that can be used to test potential functions of extracellular vesicles. Furthermore, we reveal that proteins regulating the asymmetric localization of the lipid phosphatidylethanolamine are critical for extracellular vesicle release, implicating this abundant but understudied lipid.Cells release extracellular vesicles (EVs) that mediate intercellular communication and repair damaged membranes. Despite the pleiotropic functions of EVs in vitro, their in vivo function is debated, largely because it is unclear how to induce or inhibit their formation. In particular, the mechanisms of EV release by plasma membrane budding or ectocytosis are poorly understood. We previously showed that TAT-5 phospholipid flippase activity maintains the asymmetric localization of the lipid phosphatidylethanolamine (PE) in the plasma membrane and inhibits EV budding by ectocytosis in Caenorhabditis elegans. However, no proteins that inhibit ectocytosis upstream of TAT-5 were known. Here, we identify TAT-5 regulators associated with retrograde endosomal recycling: PI3Kinase VPS-34, Beclin1 homolog BEC-1, DnaJ protein RME-8, and the uncharacterized Dopey homolog PAD-1. PI3Kinase, RME-8, and semiredundant sorting nexins are required for the plasma membrane localization of TAT-5, which is important to maintain PE asymmetry and inhibit EV release. PAD-1 does not directly regulate TAT-5 localization, but is required for the lipid flipping activity of TAT-5. PAD-1 also has roles in endosomal trafficking with the GEF-like protein MON-2, which regulates PE asymmetry and EV release redundantly with sorting nexins independent of the core retromer. Thus, in addition to uncovering redundant intracellular trafficking pathways, our study identifies additional proteins that regulate EV release. This work pinpoints TAT-5 and PE as key regulators of plasma membrane budding, further supporting the model that PE externalization drives ectocytosis.

@article{BeerE1127, abstract = {Cells must interact with their environment to survive. The lipids and proteins of the plasma membrane send and receive signals at the cell surface to respond to stimuli. When the lipid bilayer of the plasma membrane is damaged, cells release membrane-bound extracellular vesicles to repair the membrane. Cells also release signals on extracellular vesicles to communicate at a distance. Here, we identify proteins that regulate the formation of extracellular vesicles from the plasma membrane, providing additional tools to control their release that can be used to test potential functions of extracellular vesicles. Furthermore, we reveal that proteins regulating the asymmetric localization of the lipid phosphatidylethanolamine are critical for extracellular vesicle release, implicating this abundant but understudied lipid.Cells release extracellular vesicles (EVs) that mediate intercellular communication and repair damaged membranes. Despite the pleiotropic functions of EVs in vitro, their in vivo function is debated, largely because it is unclear how to induce or inhibit their formation. In particular, the mechanisms of EV release by plasma membrane budding or ectocytosis are poorly understood. We previously showed that TAT-5 phospholipid flippase activity maintains the asymmetric localization of the lipid phosphatidylethanolamine (PE) in the plasma membrane and inhibits EV budding by ectocytosis in Caenorhabditis elegans. However, no proteins that inhibit ectocytosis upstream of TAT-5 were known. Here, we identify TAT-5 regulators associated with retrograde endosomal recycling: PI3Kinase VPS-34, Beclin1 homolog BEC-1, DnaJ protein RME-8, and the uncharacterized Dopey homolog PAD-1. PI3Kinase, RME-8, and semiredundant sorting nexins are required for the plasma membrane localization of TAT-5, which is important to maintain PE asymmetry and inhibit EV release. PAD-1 does not directly regulate TAT-5 localization, but is required for the lipid flipping activity of TAT-5. PAD-1 also has roles in endosomal trafficking with the GEF-like protein MON-2, which regulates PE asymmetry and EV release redundantly with sorting nexins independent of the core retromer. Thus, in addition to uncovering redundant intracellular trafficking pathways, our study identifies additional proteins that regulate EV release. This work pinpoints TAT-5 and PE as key regulators of plasma membrane budding, further supporting the model that PE externalization drives ectocytosis.}, added-at = {2018-12-15T07:46:29.000+0100}, author = {Beer, Katharina B. and Rivas-Castillo, Jennifer and Kuhn, Kenneth and Fazeli, Gholamreza and Karmann, Birgit and Nance, Jeremy F. and Stigloher, Christian and Wehman, Ann M.}, biburl = {https://www.bibsonomy.org/bibtex/2ebe87036bb8346ee17110b6ac8ed04d7/zam}, doi = {10.1073/pnas.1714085115}, eprint = {https://www.pnas.org/content/115/6/E1127.full.pdf}, interhash = {e7d3d9345cc5d1d84f0a9e7cf4a1cc69}, intrahash = {ebe87036bb8346ee17110b6ac8ed04d7}, issn = {0027-8424}, journal = {Proceedings of the National Academy of Sciences}, keywords = {myown}, number = 6, pages = {E1127--E1136}, publisher = {National Academy of Sciences}, timestamp = {2021-02-25T18:28:59.000+0100}, title = {Extracellular vesicle budding is inhibited by redundant regulators of TAT-5 flippase localization and phospholipid asymmetry}, url = {https://www.pnas.org/content/115/6/E1127}, volume = 115, year = 2018 }