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Artificial targeting of autophagy components to mitochondria reveals both conventional and unconventional mitophagy pathways.

作者信息

Lorentzen Katharina C, Prescott Alan R, Ganley Ian G

机构信息

MRC Protein Phosphorylation and Ubiquitylation Unit, University of Dundee, Dundee, UK.

Dundee Imaging Facility, School of Life Sciences, University of Dundee, Dundee, UK.

出版信息

Autophagy. 2025 Feb;21(2):315-337. doi: 10.1080/15548627.2024.2395149. Epub 2024 Sep 8.


DOI:10.1080/15548627.2024.2395149
PMID:39177530
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC11760219/
Abstract

Macroautophagy/autophagy enables lysosomal degradation of a diverse array of intracellular material. This process is essential for normal cellular function and its dysregulation is implicated in many diseases. Given this, there is much interest in understanding autophagic mechanisms of action in order to determine how it can be best targeted therapeutically. In mitophagy, the selective degradation of mitochondria via autophagy, mitochondria first need to be primed with signals that allow the recruitment of the core autophagy machinery to drive the local formation of an autophagosome around the target mitochondrion. To determine how the recruitment of different core autophagy components can drive mitophagy, we took advantage of the -QC mitophagy assay (an outer mitochondrial membrane-localized tandem mCherry-GFP tag). By tagging autophagy proteins with an anti-mCherry (or anti-GFP) nanobody, we could recruit them to mitochondria and simultaneously monitor levels of mitophagy. We found that targeting ULK1, ATG16L1 and the different Atg8-family proteins was sufficient to induce mitophagy. Mitochondrial recruitment of ULK1 and the Atg8-family proteins induced a conventional mitophagy pathway, requiring RB1CC1/FIP200, PIK3C3/VPS34 activity and ATG5. Surprisingly, the mitophagy pathway upon recruitment of ATG16L1 proceeded independently of ATG5, although it still required RB1CC1 and PIK3C3/VPS34 activity. In this latter pathway, mitochondria were alternatively delivered to lysosomes via uptake into early endosomes. aGFP: anti-GFP nanobody; amCh: anti-mCherry nanobody; ATG: autophagy related; ATG16L1: autophagy related 16 like 1; AUTAC/AUTOTAC: autophagy-targeting chimera; BafA1: bafilomycin A; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; CCCP: carbonyl cyanide m-chlorophenylhydrazone; COX4/COX IV: cytochrome c oxidase subunit 4; DFP: deferiprone; DMSO: dimethyl sulfoxide; GABARAP: GABA type A receptor-associated protein; GABARAPL1: GABA type A receptor associated protein like 1; HSPD1/HSP60: heat shock protein family D (Hsp60) member 1; HRP: horseradish peroxidase; HTRA2/OMI: HtrA serine peptidase 2; IB: immunoblotting; IF: immunofluorescence; KO: knockout; LAMP1: lysosomal associated membrane protein 1; LIR: LC3-interacting region; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MEF: mouse embryonic fibroblast; NBR1: NBR1 autophagy cargo receptor; OMM: outer mitochondrial membrane; OPA1: OPA1 mitochondrial dynamin like GTPase; OPTN: optineurin; (D)PBS: (Dulbecco's) phosphate-buffered saline; PD: Parkinson disease; PFA: paraformaldehyde; POI: protein of interest; PtdIns3K: class III phosphatidylinositol 3-kinase; PtdIns3P: phosphatidylinositol-3-phosphate; RAB: RAB, member RAS oncogene family; RB1CC1/FIP200: RB1 inducible coiled-coil 1; SQSTM1: sequestosome 1; TAX1BP1: Tax1 binding protein 1; ULK: unc-51 like autophagy activating kinase 1; VPS: vacuolar protein sorting; WIPI: WD repeat domain, phosphoinositide interacting.

摘要
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3851/11760219/95321892e6f8/KAUP_A_2395149_F0008_C.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3851/11760219/5dc32713d8d9/KAUP_A_2395149_F0001_C.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3851/11760219/84eca5ebc8c5/KAUP_A_2395149_F0002_C.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3851/11760219/6807ebedc659/KAUP_A_2395149_F0003_C.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3851/11760219/194e3210db64/KAUP_A_2395149_F0004_C.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3851/11760219/8ace2ed7212a/KAUP_A_2395149_F0005_C.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3851/11760219/f0a855532589/KAUP_A_2395149_F0006_C.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3851/11760219/95321892e6f8/KAUP_A_2395149_F0008_C.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3851/11760219/5dc32713d8d9/KAUP_A_2395149_F0001_C.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3851/11760219/84eca5ebc8c5/KAUP_A_2395149_F0002_C.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3851/11760219/6807ebedc659/KAUP_A_2395149_F0003_C.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3851/11760219/194e3210db64/KAUP_A_2395149_F0004_C.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3851/11760219/8ace2ed7212a/KAUP_A_2395149_F0005_C.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3851/11760219/f0a855532589/KAUP_A_2395149_F0006_C.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3851/11760219/95321892e6f8/KAUP_A_2395149_F0008_C.jpg

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本文引用的文献

[1]
Tethering ATG16L1 or LC3 induces targeted autophagic degradation of protein aggregates and mitochondria.

Autophagy. 2023-11

[2]
The autophagy pathway beyond model organisms: an evolutionary perspective.

Autophagy. 2023-1

[3]
BNIP3L/NIX regulates both mitophagy and pexophagy.

EMBO J. 2022-12-15

[4]
Ispinesib as an Effective Warhead for the Design of Autophagosome-Tethering Chimeras: Discovery of Potent Degraders of Nicotinamide Phosphoribosyltransferase (NAMPT).

J Med Chem. 2022-6-9

[5]
V-ATPase is a universal regulator of LC3-associated phagocytosis and non-canonical autophagy.

J Cell Biol. 2022-6-6

[6]
The AUTOTAC chemical biology platform for targeted protein degradation via the autophagy-lysosome system.

Nat Commun. 2022-2-16

[7]
PROTAC targeted protein degraders: the past is prologue.

Nat Rev Drug Discov. 2022-3

[8]
Ulk1-dependent alternative mitophagy plays a protective role during pressure overload in the heart.

Cardiovasc Res. 2022-9-20

[9]
Activation Mechanisms of the VPS34 Complexes.

Cells. 2021-11-11

[10]
Developing potent LC3-targeting AUTAC tools for protein degradation with selective autophagy.

Chem Commun (Camb). 2021-12-7

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