TY - JOUR
T1 - Membrane characteristics tune activities of endosomal and autophagic human VPS34 complexes
AU - Ohashi, Yohei
AU - Tremel, Shirley
AU - Masson, Glenn Robert
AU - McGinney, Lauren
AU - Boulanger, Jerome
AU - Rostislavleva, Ksenia
AU - Johnson, Christopher M.
AU - Niewczas, Izabella
AU - Clark, Jonathan
AU - Williams, Roger L.
N1 - Funding Information:
We thank Aurélien Roux and Annika Hohendahl (University of Geneva) for help with assay development. We thank Olga Perisic for help with protein expression and purification, advice on assay development and discussions, Maria Daly for help with exploring flow cytometry-based assays, Lufei Zhang for purifying VPS34 alone, Sarah Maslen and Mark Skehel for assistance with HDX-MS and Conrad Weichbrodt of Nanion Technologies for advice on preparation of GUVs. The work was supported by the MRC (File reference number MC_U105184308 to RLW) and Cancer Research UK (Programme grant C14801/A21211 to RLW). ST was supported by an MRC studentship. GRM was supported by AstraZeneca/LMB Blue Sky Initiative [MC-A024-5PF9L] and by a Henslow Research Fellowship from The Cambridge Philosophical Society and St Catharine’s College, Cambridge.
Funding Information:
We thank Aure?lien Roux and Annika Hohendahl (University of Geneva) for help with assay development. We thank Olga Perisic for help with protein expression and purification, advice on assay development and discussions, Maria Daly for help with exploring flow cytometry-based assays, Lufei Zhang for purifying VPS34 alone, Sarah Maslen and Mark Skehel for assistance with HDX-MS and Conrad Weichbrodt of Nanion Technologies for advice on preparation of GUVs. The work was supported by the MRC (File reference number MC_U105184308 to RLW) and Cancer Research UK (Programme grant C14801/A21211 to RLW). ST was supported by an MRC studentship. GRM was supported by AstraZeneca/LMB Blue Sky Initiative [MC-A024-5PF9L] and by a Henslow Research Fellowship from The Cambridge Philosophical Society and St Catharine?s College, Cambridge. Medical Research Council MC_U105184308 Roger L Williams, Cancer Research UK C14801/A21211 Roger L Williams, Medical Research Council MC-A024-5PF9L Roger L Williams.
Publisher Copyright:
© Ohashi et al.
Copyright:
Copyright 2020 Elsevier B.V., All rights reserved.
PY - 2020/6/30
Y1 - 2020/6/30
N2 - The lipid kinase VPS34 orchestrates diverse processes, including autophagy, endocytic sorting, phagocytosis, anabolic responses and cell division. VPS34 forms various complexes that help adapt it to specific pathways, with complexes I and II being the most prominent ones. We found that physicochemical properties of membranes strongly modulate VPS34 activity. Greater unsaturation of both substrate and non-substrate lipids, negative charge and curvature activate VPS34 complexes, adapting them to their cellular compartments. Hydrogen/deuterium exchange mass spectrometry (HDX-MS) of complexes I and II on membranes elucidated structural determinants that enable them to bind membranes. Among these are the Barkor/ATG14L autophagosome targeting sequence (BATS), which makes autophagy-specific complex I more active than the endocytic complex II, and the Beclin1 BARA domain. Interestingly, even though Beclin1 BARA is common to both complexes, its membrane-interacting loops are critical for complex II, but have only a minor role for complex I.
AB - The lipid kinase VPS34 orchestrates diverse processes, including autophagy, endocytic sorting, phagocytosis, anabolic responses and cell division. VPS34 forms various complexes that help adapt it to specific pathways, with complexes I and II being the most prominent ones. We found that physicochemical properties of membranes strongly modulate VPS34 activity. Greater unsaturation of both substrate and non-substrate lipids, negative charge and curvature activate VPS34 complexes, adapting them to their cellular compartments. Hydrogen/deuterium exchange mass spectrometry (HDX-MS) of complexes I and II on membranes elucidated structural determinants that enable them to bind membranes. Among these are the Barkor/ATG14L autophagosome targeting sequence (BATS), which makes autophagy-specific complex I more active than the endocytic complex II, and the Beclin1 BARA domain. Interestingly, even though Beclin1 BARA is common to both complexes, its membrane-interacting loops are critical for complex II, but have only a minor role for complex I.
UR - http://www.scopus.com/inward/record.url?scp=85087412986&partnerID=8YFLogxK
U2 - 10.7554/eLife.58281
DO - 10.7554/eLife.58281
M3 - Article
C2 - 32602837
AN - SCOPUS:85087412986
SN - 2050-084X
VL - 9
SP - 1
EP - 28
JO - eLife
JF - eLife
M1 - e58281
ER -
