Universität zu Köln

Kallabis, Sebastian, Abraham, Lena, Mueller, Stefan, Dzialas, Verena, Tuerk, Clara, Wiederstein, Janica Lea, Bock, Theresa, Nolte, Hendrik, Nogara, Leonardo, Blaauw, Bert, Braun, Thomas and Krueger, Marcus ORCID: 0000-0003-2008-4582 (2020). High-throughput proteomics fiber typing (ProFiT) for comprehensive characterization of single skeletal muscle fibers. Skeletal Muscle, 10 (1). LONDON: BMC. ISSN 2044-5040

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Abstract

Background Skeletal muscles are composed of a heterogeneous collection of fiber types with different physiological adaption in response to a stimulus and disease-related conditions. Each fiber has a specific molecular expression of myosin heavy chain molecules (MyHC). So far, MyHCs are currently the best marker proteins for characterization of individual fiber types, and several proteome profiling studies have helped to dissect the molecular signature of whole muscles and individual fibers. Methods Herein, we describe a mass spectrometric workflow to measure skeletal muscle fiber type-specific proteomes. To bypass the limited quantities of protein in single fibers, we developed a Proteomics high-throughput fiber typing (ProFiT) approach enabling profiling of MyHC in single fibers. Aliquots of protein extracts from separated muscle fibers were subjected to capillary LC-MS gradients to profile MyHC isoforms in a 96-well format. Muscle fibers with the same MyHC protein expression were pooled and subjected to proteomic, pulsed-SILAC, and phosphoproteomic analysis. Results Our fiber type-specific quantitative proteome analysis confirmed the distribution of fiber types in the soleus muscle, substantiates metabolic adaptions in oxidative and glycolytic fibers, and highlighted significant differences between the proteomes of type IIb fibers from different muscle groups, including a differential expression of desmin and actinin-3. A detailed map of the Lys-6 incorporation rates in muscle fibers showed an increased turnover of slow fibers compared to fast fibers. In addition, labeling of mitochondrial respiratory chain complexes revealed a broad range of Lys-6 incorporation rates, depending on the localization of the subunits within distinct complexes. Conclusion Overall, the ProFiT approach provides a versatile tool to rapidly characterize muscle fibers and obtain fiber-specific proteomes for different muscle groups.

Item Type:	Journal Article
Creators:	Creators Email ORCID ORCID Put Code Kallabis, Sebastian UNSPECIFIED UNSPECIFIED UNSPECIFIED Abraham, Lena UNSPECIFIED UNSPECIFIED UNSPECIFIED Mueller, Stefan UNSPECIFIED UNSPECIFIED UNSPECIFIED Dzialas, Verena UNSPECIFIED UNSPECIFIED UNSPECIFIED Tuerk, Clara UNSPECIFIED UNSPECIFIED UNSPECIFIED Wiederstein, Janica Lea UNSPECIFIED UNSPECIFIED UNSPECIFIED Bock, Theresa UNSPECIFIED UNSPECIFIED UNSPECIFIED Nolte, Hendrik UNSPECIFIED UNSPECIFIED UNSPECIFIED Nogara, Leonardo UNSPECIFIED UNSPECIFIED UNSPECIFIED Blaauw, Bert UNSPECIFIED UNSPECIFIED UNSPECIFIED Braun, Thomas UNSPECIFIED UNSPECIFIED UNSPECIFIED Krueger, Marcus UNSPECIFIED orcid.org/0000-0003-2008-4582 UNSPECIFIED
URN:	urn:nbn:de:hbz:38-340417
DOI:	10.1186/s13395-020-00226-5
Journal or Publication Title:	Skeletal Muscle
Volume:	10
Number:	1
Date:	2020
Publisher:	BMC
Place of Publication:	LONDON
ISSN:	2044-5040
Language:	English
Faculty:	Faculty of Mathematics and Natural Sciences
Divisions:	Faculty of Mathematics and Natural Sciences > Department of Biology > Institute for Genetics
Subjects:	no entry
Uncontrolled Keywords:	Keywords Language LIGHT-CHAIN PHOSPHORYLATION; COMPLEX I; ENCODED SUBUNITS; GENE-EXPRESSION; MYOSIN ISOFORMS; MITOCHONDRIAL; TURNOVER; ACTIVATION; MECHANISM; DYNAMICS Multiple languages Cell Biology Multiple languages
Refereed:	Yes
URI:	http://kups.ub.uni-koeln.de/id/eprint/34041