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Secretory autophagy mediates mitochondria clearance in the absence of the mATG8-conjugation machinery

ABSTRACT: PINK1-Parkin mediated mitophagy, a selective form of autophagy, represents an important mechanism in mitochondrial quality control (MQC) via clearance of damaged mitochondria. Although it is well known that the conjugation of mammalian ATG8s to phosphatidylethanolamine (PE) is a key step in autophagy, its role in mitophagy remains controversial. In this study, we attempted to clarify the role of the mATG8-conjugation system in mitophagy by generating knockouts of the mATG8-conjugation machinery. Unexpectedly, we show that mitochondria could still be cleared in the absence of mATG8 lipidation, in a process that was found to be independent of lysosomal degradation. Instead, mitochondria were found to be cleared via a secretory autophagy pathway, resulting in the extracellular release of mitochondria, in a process we defined as autophagic secretion of mitochondria (ASM). Functionally, increased ASM promoted the activation of the innate immune cGAS-STING pathway in recipient cells. Overall, data from this study reveal ASM as a novel mechanism in MQC, especially when the cellular mATG8-conjugation machinery is dysfunctional.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Line Cell

DISEASE(S): Cervical Cancer

SUBMITTER: Dennis Kappei

LAB HEAD: Dennis Kappei

PROVIDER: PXD025356 | Pride | 2022-06-03

REPOSITORIES: Pride

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ATG conjugation system -dependent and -independent mechanisms underlie lysosomal stress-induced TFEB regulation

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mTOR hyperactivation in Down Syndrome mediates deficits in autophagy induction, autophagosome formation, and mitophagy

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2019-06-22 | GSE126910 | GEO

The LC3 conjugation machinery specifies the loading of RNA-binding proteins into extracellular vesicles (smRNA-seq dataset)

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The LC3 conjugation machinery specifies the loading of RNA-binding proteins into extracellular vesicles (lrgRNA-seq dataset)

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PARK15/FBXO7 is dispensable for PINK1/Parkin mitophagy in iNeurons and HeLa cell systems

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2023-06-19 | PXD037797 | Pride

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Project description:Horizontal gene transfer (HGT) is the major mechanism responsible for spread of antibiotic resistance. Antibiotic treatment has been suggested to promote HGT, either by directly affecting the conjugation process itself or by selecting for conjugations subsequent to DNA transfer. However, recent research suggests that the effect of antibiotic treatment on plasmid conjugation frequencies, and hence the spread of resistance plasmids, may have been overestimated. We addressed the question by quantifying transfer proteins and conjugation frequencies of a blaCTX-M-1 encoding IncI1 resistance plasmid in Escherichia coli MG1655 in the presence and absence of therapeutically relevant concentrations of cefotaxime (CTX). Analysis of the proteome by iTRAQ labeling and liquid chromatography tandem mass spectrometry revealed that Tra proteins were significantly up regulated in the presence of CTX. The up-regulation of the transfer machinery was confirmed at the transcriptional level for five selected genes. The CTX treatment did not cause induction of the SOS39 response as revealed by absence of significantly regulated SOS associated proteins in the proteome and no significant up-regulation of recA and sfiA genes. The frequency of plasmid conjugation, measured in an antibiotic free environment, increased significantly when the donor was pre-grown in broth containing CTX compared to growth without this drug, regardless of whether blaCTX-M-1 was located on the plasmid or in trans on the chromosome. The results shows that antibiotic treatment can affect expression of a plasmid conjugation machinery and subsequent DNA transfer.

2017-11-21 | PXD006679 | Pride

Phosphosite mapping on PI3Kc1 by ULK1 and TBK1 kinases.

Project description:Autophagy is a highly conserved catabolic process which degrades and recycles intracellular components. To accomplish this task the surplus material (cargo) is sequestered in a double membranous vesicle termed autophagosome. To initiate the formation of autophagosome autophagy machinery is recruited to cargo in a controlled manner. One of the first factors recruited to damaged mitochondria during mitophagy (selective removal of damaged mitochondria by autophagy) are two protein kinases, ULK1/2 complex and TBK1. They both promote a subsequent recruitment of the class III phosphatidylinositol 3-kinase complex I (PI3Kc1 complex) which function is to generate the lipid phosphatidylinositol 3-phosphate (PI(3)P) on phagophore membranes. Both ULK1 and TBK1 were reported to phosphorylate several autophagy factors. Here, we probed phosphorylation of PI3Kc1 by ULK1 and TBK1 to gain a full picture of the phosphosites map of PI3Kc phosphorylated by both protein kinases. Recombinant PI3Kc1 was incubated with recombinant ULK1 complex or TBK1 in the presence of ATP and MgCl2. As a negative control, the same protein mixtures were incubated in a kinase buffer lacking MgCl2 and ATP. Samples were separated by SDS-PAGE and stained with Coomassie. The bands for each subunit of the PI3KC3-C1 complex (VPS15, VPS34, ATG14, and Beclin1) were extracted from the gel and submitted for mass spectrometry analysis.

2023-05-08 | PXD039663 | Pride

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2022-01-17 | PXD027586 | Pride

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