Dataset Information

IP-MS of human YKT6 upon mutation of possible ULK1-phosphorylation sites

ABSTRACT: Autophagy is a catabolic process during which cytosolic material is enwrapped in a newly formed double membrane structure called the autophagosome, and subsequently targeted for degradation in the lytic compartment of the cell. The fusion of autophagosomes with the lytic compartment is a tightly regulated step and involves membrane-bound SNARE proteins. These play a crucial role as they promote lipid mixing and fusion of the opposing membranes. Among the SNARE proteins implicated in autophagy, the essential SNARE protein YKT6 is the only evolutionary conserved SNARE protein from yeast to human. Alterations in YKT6 function, in both mammalian cells and nematodes, produces early and late autophagy defects that result in reduced survival. Moreover, mammalian autophagosomal YKT6 is phospho-regulated by the ULK1 kinase, preventing premature bundling with the lysosomal SNARE proteins and autophagosome-lysosome fusion. Together, our findings reveal that timely regulation of theYKT6 phosphostatus is crucial throughout autophagy progression and cell survival.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Epithelial Cell

SUBMITTER: Georg Tascher

LAB HEAD: Claudine Kraft

PROVIDER: PXD038148 | Pride | 2023-03-02

REPOSITORIES: pride

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Dataset's files

Source:

			Action	DRS
	20211119_GT_PSM_YKT6_IP_CTRL_1_211119172237.raw	Raw
	20211119_GT_PSM_YKT6_IP_CTRL_2.raw	Raw
	20211119_GT_PSM_YKT6_IP_CTRL_3.raw	Raw
	20211119_GT_PSM_YKT6_IP_KO_1.raw	Raw
	20211119_GT_PSM_YKT6_IP_KO_2.raw	Raw

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Publications

ULK1-mediated phosphorylation regulates the conserved role of YKT6 in autophagy.

Sánchez-Martín Pablo P Kriegenburg Franziska F Alves Ludovico L Adam Julius J Elsaesser Jana J Babic Riccardo R Mancilla Hector H Licheva Mariya M Tascher Georg G Münch Christian C Eimer Stefan S Kraft Claudine C

Journal of cell science 20230209 3

Autophagy is a catabolic process during which cytosolic material is enwrapped in a newly formed double-membrane structure called the autophagosome, and subsequently targeted for degradation in the lytic compartment of the cell. The fusion of autophagosomes with the lytic compartment is a tightly regulated step and involves membrane-bound SNARE proteins. These play a crucial role as they promote lipid mixing and fusion of the opposing membranes. Among the SNARE proteins implicated in autophagy, t ...[more]

PMID: 36644903

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Project description:Removal of damaged organelles via the process of selective autophagy constitutes a major form of cellular quality control. Damaged organelles are recognized by a dedicated surveillance machinery, leading to the assembly of an autophagosome around the damaged organelle, prior to fusion with the degradative lysosomal compartment. Lysosomes themselves are also prone to damage and are degraded through the process of lysophagy. While early steps involve recognition of ruptured lysosomal membranes by glycan-binding Galectins and ubiquitylation of transmembrane lysosomal proteins, many steps in the process, and their inter-relationships, remain poorly understood, including the role and identity of cargo receptors required for completion of lysophagy. Here, we employ quantitative organelle capture and proximity biotinylation proteomics of autophagy adaptors, cargo receptors, and Galectins in response to acute lysosomal damage, thereby revealing the landscape of lysosomal proteome remodeling during lysophagy. Among proteins dynamically recruited to damaged lysosomes were ubiquitin-binding autophagic cargo receptors. Using newly developed lysophagic flux reporters including Lyso-Keima, we demonstrate that TAX1BP1, together with its associated kinase TBK1, are both necessary and sufficient to promote lysophagic flux in both Hela cells and induced neurons (iNeurons). While the related receptor OPTN can drive damage-dependent lysophagy when overexpressed, cells lacking either OPTN or CALCOCO2 still maintain significant lysophagic flux in HeLa cells. Mechanistically, TAX1BP1-driven lysophagy requires its N-terminal SKICH domain, which binds both TBK1 and the autophagy regulatory factor RB1CC1, and requires upstream ubiquitylation events for efficient recruitment and lysophagic flux. These results identify TAX1BP1 as a central component in the lysophagy pathway and provide a proteomic resource for future studies of the lysophagy process.