PXD074963

PXD074963 is an original dataset announced via ProteomeXchange.

Title	A GABARAP−PtdIns3K-C1 positive feedback loop at the heart of the phagophore nucleation
Description	Macroautophagy/autophagy is a cellular process allowing the degradation of intracellular components during starvation. In mammalian cells, autophagosomes can reach over 1000 nm within 30 min after triggering starvation, but how such substantial amounts of membranes can be synthesized within a brief time remains elusive. A protein complex central to the phagophore initiation is the lipid kinase PIK3C3-Complex 1 (PtdIns3K-C1), which produces phosphatidylinositol-3-phosphate (PtdIns3P). PtdIns3P recruits a variety of downstream proteins, among which are the mammalian ATG8 (mATG8) family proteins which are coupled to the lipids of the phagophore. mATG8s were previously suggested to interact with peptides derived from PtdIns3K-C1, but potential functional implications on the full-complex assembly remained unknown. Here we show that recombinant membrane-coupled mATG8s bind to and potently activate PtdIns3K-C1, with GABARAP being the most potent activator among all mATG8s. By a combination of activity assays, cryo-electronic microscopy (cryo-EM), structural mass spectrometry and mutagenesis, we show that GABARAP binds two sites in PtdIns3K-C1 out of the three that were initially proposed based on peptide binding, with one site showing an atypical bipartite interaction with the mATG8. We also confirm both sites are essential for GABARAP to activate PtdIns3K-C1. We propose this activation of PtdIns3K-C1 by GABARAP forms an unexpected positive feedback loop in cells, whereby basal levels of PtdIns3P production by PtdIns3K-C1 indirectly recruit GABARAP, which in turn helps PtdIns3K-C1 produce more PtdIns3P. This mechanism would be central for autophagosome biogenesis, where enlarged membranes need to be synthesized within a brief period.
HostingRepository	PRIDE
AnnounceDate	2026-07-02
AnnouncementXML	Submission_2026-07-02_05:49:56.255.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Maxime Bourguet
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: NEWT:9606;
ModificationList	No PTMs are included in the dataset
Instrument	Synapt MS

Revision	Datetime	Status	ChangeLog Entry
0	2026-02-26 16:42:55	ID requested
⏵ 1	2026-07-02 05:49:57	announced

Dataset with its publication pending

submitter keyword: autophagy

cryo-EM

GABARAP

HDX-MS

LC3

positive feedback loop

XL-MS

Roger Lee Williams
contact affiliation	MRC - Laboratory of Molecular Biology
contact email	rlw@mrc-lmb.cam.ac.uk
lab head
Maxime Bourguet
contact affiliation	MRC Laboratory of Molecular Biology
contact email	mbourguet@mrc-lmb.cam.ac.uk
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/07/PXD074963

PRIDE project URI

• PRIDE
  1. PXD074963
     1. Label: PRIDE project
     2. Name: A GABARAP−PtdIns3K-C1 positive feedback loop at the heart of the phagophore nucleation