TNIP1 inhibits selective autophagy via bipartite interaction with LC3/GABARAP and TAX1BP1

Creators: Le Guerroué, François; Bunker, Eric N.; Rosencrans, William M.; Nguyen, Jack T.; Basar, Mohammed A.; Werner, Achim; Chou, Tsui-Fen¹; Wang, Chunxin; Youle, Richard J.

1. California Institute of Technology

Abstract

Mitophagy is a form of selective autophagy that disposes of superfluous and potentially damage-inducing organelles in a tightly controlled manner. While the machinery involved in mitophagy induction is well known, the regulation of the components is less clear. Here, we demonstrate that TNIP1 knockout in HeLa cells accelerates mitophagy rates and that ectopic TNIP1 negatively regulates the rate of mitophagy. These functions of TNIP1 depend on an evolutionarily conserved LIR motif as well as an AHD3 domain, which are required for binding to the LC3/GABARAP family of proteins and the autophagy receptor TAX1BP1, respectively. We further show that phosphorylation appears to regulate its association with the ULK1 complex member FIP200, allowing TNIP1 to compete with autophagy receptors, which provides a molecular rationale for its inhibitory function during mitophagy. Taken together, our findings describe TNIP1 as a negative regulator of mitophagy that acts at the early steps of autophagosome biogenesis.

Copyright and License

Published by Elsevier Under an Elsevier user license.

Acknowledgement

The authors would like to thank the laboratory of Michael Lazarou (Monash Biomedicine Discovery Institute) for sharing the LC3/GABARAP KO cells. We would also like to thank all other members of the Youle lab for sharing cell lines and reagents as well as discussions and advice. We further thank Dr Yan Wang from the National Institute of Dental and Craniofacial Research Mass Spectrometry Facility (ZIA DE00075) for performing mass spectrometry analysis. FACS data were acquired using the NINDS facility under the supervision of Dr. Dragan Maric. This work was supported by the Intramural Program of the NINDS and the NIDCR, NIH.

Contributions

E.N.B. analyzed live cell imaging data, ubiquitin foci, and quantified immunofluorescence data. W.M.R. and J.T.N. performed and analyzed FP and DLS experiments. F.L.G. conceived the study, performed all other experiments, and analyzed and interpreted all other data. M.A.B. produced and purified TNIP1 recombinant protein. A.W. performed MS and analyzed MS data. C.W. provided reagents and cell lines. F.L.G. and R.J.Y. wrote the manuscript with input from all authors. R.J.Y., A.W., and T.-F.C. supervised the project and acquired funds.

Conflict of Interest

The authors declare no financial conflicts and assure that this manuscript is original and has not been published nor is currently under consideration for publication elsewhere.

Data Availability

igh content imaging data is deposited at BioImage Archive: https://www.ebi.ac.uk/biostudies/BioImages/studies/S-BIAD619. Original Western blot data, microscopy images, raw FACS data, raw MS data, raw FP data, and code used in this paper have been deposited at Mendeley Data and are publicly available as of the date of publication at https://doi.org/10.17632/jh7h5cx4yh.1. MS data are deposited at the MassIVE repository under the accession number MassIVE: MSV000091090.
o new code has been generated for this study.
Any additional information required to reanalyze the data reported in this paper is available from the lead contact upon request.

Additional Information

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