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Lookup NU author(s): Manuel Banzhaf
This work is licensed under a Creative Commons Attribution 4.0 International License (CC BY 4.0).
© 2020 The Authors. The FASEB Journal published by Wiley Periodicals, Inc. on behalf of Federation of American Societies for Experimental Biology. Mitophagy is a key process regulating mitochondrial quality control. Several mechanisms have been proposed to regulate mitophagy, but these have mostly been studied using stably expressed non-native proteins in immortalized cell lines. In skeletal muscle, mitophagy and its molecular mechanisms require more thorough investigation. To measure mitophagy directly, we generated a stable skeletal muscle C2C12 cell line, expressing a mitophagy reporter construct (mCherry-green fluorescence protein-mtFIS1101-152). Here, we report that both carbonyl cyanide m-chlorophenyl hydrazone (CCCP) treatment and adenosine monophosphate activated protein kinase (AMPK) activation by 991 promote mitochondrial fission via phosphorylation of MFF and induce mitophagy by ~20%. Upon CCCP treatment, but not 991, ubiquitin phosphorylation, a read-out of PTEN-induced kinase 1 (PINK1) activity, and Parkin E3 ligase activity toward CDGSH iron sulfur domain 1 (CISD1) were increased. Although the PINK1-Parkin signaling pathway is active in response to CCCP treatment, we observed no change in markers of mitochondrial protein content. Interestingly, our data shows that TANK-binding kinase 1 (TBK1) phosphorylation is increased after both CCCP and 991 treatments, suggesting TBK1 activation to be independent of both PINK1 and Parkin. Finally, we confirmed in non-muscle cell lines that TBK1 phosphorylation occurs in the absence of PINK1 and is regulated by AMPK-dependent signaling. Thus, AMPK activation promotes mitophagy by enhancing mitochondrial fission (via MFF phosphorylation) and autophagosomal engulfment (via TBK1 activation) in a PINK1-Parkin independent manner.
Author(s): Seabright AP, Fine NHF, Barlow JP, Lord SO, Musa I, Gray A, Bryant JA, Banzhaf M, Lavery GG, Hardie DG, Hodson DJ, Philp A, Lai Y-C
Publication type: Article
Publication status: Published
Journal: FASEB Journal
Year: 2020
Volume: 34
Issue: 5
Pages: 6284-6301
Print publication date: 01/05/2020
Online publication date: 22/03/2020
Acceptance date: 28/02/2020
Date deposited: 14/08/2023
ISSN (print): 0892-6638
ISSN (electronic): 1530-6860
Publisher: John Wiley and Sons Inc.
URL: https://doi.org/10.1096/fj.201903051R
DOI: 10.1096/fj.201903051R
PubMed id: 32201986
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