Universität zu Köln

Hofsetz, Eduard ORCID: 0000-0001-5830-4566, Demir, Fatih ORCID: 0000-0002-5744-0205, Szczepanowska, Karolina ORCID: 0000-0003-4689-2350, Kukat, Alexandra, Kizhakkedathu, Jayachandran N., Trifunovic, Aleksandra and Huesgen, Pitter F. (2020). The Mouse Heart Mitochondria N Terminome Provides Insights into ClpXP-Mediated Proteolysis. Mol. Cell. Proteomics, 19 (8). S. 1330 - 1346. ROCKVILLE: AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC. ISSN 1535-9484

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Abstract

The mammalian mitochondrial proteome consists of more than 1100 annotated proteins and their proteostasis is regulated by only a few ATP-dependent protease complexes. Technical advances in protein mass spectrometry allowed for detailed description of the mitoproteome from different species and tissues and their changes under specific conditions. However, protease-substrate relations within mitochondria are still poorly understood. Here, we combined Terminal Amine Isotope Labeling of Substrates (TAILS) N termini profiling of heart mitochondria proteomes isolated from wild type and Clpp(-/-) mice with a classical substrate-trapping screen using FLAG-tagged proteolytically active and inactive CLPP variants to identify new ClpXP substrates in mammalian mitochondria. Using TAILS, we identified N termini of more than 200 mitochondrial proteins. Expected N termini confirmed sequence determinants for mitochondrial targeting signal (MTS) cleavage and subsequent N-terminal processing after import, but the majority were protease-generated neo-N termini mapping to positions within the proteins. Quantitative comparison revealed widespread changes in protein processing patterns, including both strong increases or decreases in the abundance of specific neo-N termini, as well as an overall increase in the abundance of protease-generated neo-N termini in CLPP-deficient mitochondria that indicated altered mitochondrial proteostasis. Based on the combination of altered processing patterns, protein accumulation and stabilization in CLPP-deficient mice and interaction with CLPP, we identified OAT, HSPA9 and POLDIP2 and as novel bona fide ClpXP substrates. Finally, we propose that ClpXP participates in the cooperative degradation of UQCRC1. Together, our data provide the first landscape of the heart mitochondria N terminome and give further insights into regulatory and assisted proteolysis mediated by ClpXP.

Item Type:	Journal Article
Creators:	Creators Email ORCID ORCID Put Code Hofsetz, Eduard UNSPECIFIED orcid.org/0000-0001-5830-4566 UNSPECIFIED Demir, Fatih UNSPECIFIED orcid.org/0000-0002-5744-0205 UNSPECIFIED Szczepanowska, Karolina UNSPECIFIED orcid.org/0000-0003-4689-2350 UNSPECIFIED Kukat, Alexandra UNSPECIFIED UNSPECIFIED UNSPECIFIED Kizhakkedathu, Jayachandran N. UNSPECIFIED UNSPECIFIED UNSPECIFIED Trifunovic, Aleksandra UNSPECIFIED UNSPECIFIED UNSPECIFIED Huesgen, Pitter F. UNSPECIFIED UNSPECIFIED UNSPECIFIED
URN:	urn:nbn:de:hbz:38-324811
DOI:	10.1074/mcp.RA120.002082
Journal or Publication Title:	Mol. Cell. Proteomics
Volume:	19
Number:	8
Page Range:	S. 1330 - 1346
Date:	2020
Publisher:	AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
Place of Publication:	ROCKVILLE
ISSN:	1535-9484
Language:	English
Faculty:	Unspecified
Divisions:	Unspecified
Subjects:	no entry
Uncontrolled Keywords:	Keywords Language COMPUTATIONAL PLATFORM; PROTEASE REVEALS; IN-VIVO; DEGRADATION; LON; RECOGNITION; PROTEINS; PHOSPHORYLATION; IDENTIFICATION; DISCOVERY Multiple languages Biochemical Research Methods Multiple languages
URI:	http://kups.ub.uni-koeln.de/id/eprint/32481