biostudies-literatureNguyen TTHoward Hughes Medical InstituteNational Institutes of Healthe84279https://www.ebi.ac.uk/biostudies/studies/S-EPMC9725753biostudies-literatureUnknown11Mitochondria are dynamic organelles that undergo cycles of fission and fusion at a unified platform defined by endoplasmic reticulum (ER)-mitochondria membrane contact sites (MCSs). These MCSs or nodes co-localize fission and fusion machinery. We set out to identify how ER-associated mitochondrial nodes can regulate both fission and fusion machinery assembly. We have used a promiscuous biotin ligase linked to the fusion machinery, Mfn1, and proteomics to identify an ER membrane protein, ABHD16A, as a major regulator of node formation. In the absence of ABHD16A, fission and fusion machineries fail to recruit to ER-associated mitochondrial nodes, and fission and fusion rates are significantly reduced. ABHD16A contains an acyltransferase motif and an α/β hydrolase domain, and point mutations in critical residues of these regions fail to rescue the formation of ER-associated mitochondrial hot spots. These data suggest a mechanism whereby ABHD16A functions by altering phospholipid composition at ER-mitochondria MCSs. Our data present the first example of an ER membrane protein that regulates the recruitment of both fission and fusion machineries to mitochondria.eLifeAn ER phospholipid hydrolase drives ER-associated mitochondrial constriction for fission and fusion.PMC9725753T32 Training Grants GM008759 and GM142607Nguyen TTVoeltz GKfalseAn ER phospholipid hydrolase drives ER-associated mitochondrial constriction for fission and fusion.Mitochondria are dynamic organelles that undergo cycles of fission and fusion at a unified platform defined by endoplasmic reticulum (ER)-mitochondria membrane contact sites (MCSs). These MCSs or nodes co-localize fission and fusion machinery. We set out to identify how ER-associated mitochondrial nodes can regulate both fission and fusion machinery assembly. We have used a promiscuous biotin ligase linked to the fusion machinery, Mfn1, and proteomics to identify an ER membrane protein, ABHD16A, as a major regulator of node formation. In the absence of ABHD16A, fission and fusion machineries fail to recruit to ER-associated mitochondrial nodes, and fission and fusion rates are significantly reduced. ABHD16A contains an acyltransferase motif and an α/β hydrolase domain, and point mutations in critical residues of these regions fail to rescue the formation of ER-associated mitochondrial hot spots. These data suggest a mechanism whereby ABHD16A functions by altering phospholipid composition at ER-mitochondria MCSs. Our data present the first example of an ER membrane protein that regulates the recruitment of both fission and fusion machineries to mitochondria.2022-01-01T00:00:00Z2022 Nov2025-04-22T00:38:45.832Z2025-04-05T19:35:37.81ZS-EPMC97257533644854110.7554/eLife.84279