Secretome Analysis of Lipid-Induced Insulin Resistance in Skeletal Muscle Cells by a Combined Experimental and Bioinformatics Workflow

Skeletal muscle has emerged as an important secretory organ that produces so-called myokines, regulating energy metabolism via autocrine, paracrine, and endocrine actions; however, the nature and extent of the muscle secretome has not been fully elucidated. Mass spectrometry (MS)-based proteomics, i...

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Veröffentlicht in:Journal of proteome research Jg. 14; H. 11; S. 4885 - 4895
Hauptverfasser: Deshmukh, Atul S, Cox, Juergen, Jensen, Lars Juhl, Meissner, Felix, Mann, Matthias
Format: Journal Article
Sprache:Englisch
Veröffentlicht: United States American Chemical Society 06.11.2015
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ISSN:1535-3893, 1535-3907, 1535-3907
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Abstract Skeletal muscle has emerged as an important secretory organ that produces so-called myokines, regulating energy metabolism via autocrine, paracrine, and endocrine actions; however, the nature and extent of the muscle secretome has not been fully elucidated. Mass spectrometry (MS)-based proteomics, in principle, allows an unbiased and comprehensive analysis of cellular secretomes; however, the distinction of bona fide secreted proteins from proteins released upon lysis of a small fraction of dying cells remains challenging. Here we applied highly sensitive MS and streamlined bioinformatics to analyze the secretome of lipid-induced insulin-resistant skeletal muscle cells. Our workflow identified 1073 putative secreted proteins including 32 growth factors, 25 cytokines, and 29 metalloproteinases. In addition to previously reported proteins, we report hundreds of novel ones. Intriguingly, ∼40% of the secreted proteins were regulated under insulin-resistant conditions, including a protein family with signal peptide and EGF-like domain structure that had not yet been associated with insulin resistance. Finally, we report that secretion of IGF and IGF-binding proteins was down-regulated under insulin-resistant conditions. Our study demonstrates an efficient combined experimental and bioinformatics workflow to identify putative secreted proteins from insulin-resistant skeletal muscle cells, which could easily be adapted to other cellular models.
AbstractList Skeletal muscle has emerged as an important secretory organ that produces so-called myokines, regulating energy metabolism via autocrine, paracrine, and endocrine actions; however, the nature and extent of the muscle secretome has not been fully elucidated. Mass spectrometry (MS)-based proteomics, in principle, allows an unbiased and comprehensive analysis of cellular secretomes; however, the distinction of bona fide secreted proteins from proteins released upon lysis of a small fraction of dying cells remains challenging. Here we applied highly sensitive MS and streamlined bioinformatics to analyze the secretome of lipid-induced insulin-resistant skeletal muscle cells. Our workflow identified 1073 putative secreted proteins including 32 growth factors, 25 cytokines, and 29 metalloproteinases. In addition to previously reported proteins, we report hundreds of novel ones. Intriguingly, ∼40% of the secreted proteins were regulated under insulin-resistant conditions, including a protein family with signal peptide and EGF-like domain structure that had not yet been associated with insulin resistance. Finally, we report that secretion of IGF and IGF-binding proteins was down-regulated under insulin-resistant conditions. Our study demonstrates an efficient combined experimental and bioinformatics workflow to identify putative secreted proteins from insulin-resistant skeletal muscle cells, which could easily be adapted to other cellular models.
Skeletal muscle has emerged as an important secretory organ that produces so-called myokines, regulating energy metabolism via autocrine, paracrine, and endocrine actions; however, the nature and extent of the muscle secretome has not been fully elucidated. Mass spectrometry (MS)-based proteomics, in principle, allows an unbiased and comprehensive analysis of cellular secretomes; however, the distinction of bona fide secreted proteins from proteins released upon lysis of a small fraction of dying cells remains challenging. Here we applied highly sensitive MS and streamlined bioinformatics to analyze the secretome of lipid-induced insulin-resistant skeletal muscle cells. Our workflow identified 1073 putative secreted proteins including 32 growth factors, 25 cytokines, and 29 metalloproteinases. In addition to previously reported proteins, we report hundreds of novel ones. Intriguingly, ∼40% of the secreted proteins were regulated under insulin-resistant conditions, including a protein family with signal peptide and EGF-like domain structure that had not yet been associated with insulin resistance. Finally, we report that secretion of IGF and IGF-binding proteins was down-regulated under insulin-resistant conditions. Our study demonstrates an efficient combined experimental and bioinformatics workflow to identify putative secreted proteins from insulin-resistant skeletal muscle cells, which could easily be adapted to other cellular models.Skeletal muscle has emerged as an important secretory organ that produces so-called myokines, regulating energy metabolism via autocrine, paracrine, and endocrine actions; however, the nature and extent of the muscle secretome has not been fully elucidated. Mass spectrometry (MS)-based proteomics, in principle, allows an unbiased and comprehensive analysis of cellular secretomes; however, the distinction of bona fide secreted proteins from proteins released upon lysis of a small fraction of dying cells remains challenging. Here we applied highly sensitive MS and streamlined bioinformatics to analyze the secretome of lipid-induced insulin-resistant skeletal muscle cells. Our workflow identified 1073 putative secreted proteins including 32 growth factors, 25 cytokines, and 29 metalloproteinases. In addition to previously reported proteins, we report hundreds of novel ones. Intriguingly, ∼40% of the secreted proteins were regulated under insulin-resistant conditions, including a protein family with signal peptide and EGF-like domain structure that had not yet been associated with insulin resistance. Finally, we report that secretion of IGF and IGF-binding proteins was down-regulated under insulin-resistant conditions. Our study demonstrates an efficient combined experimental and bioinformatics workflow to identify putative secreted proteins from insulin-resistant skeletal muscle cells, which could easily be adapted to other cellular models.
Author Deshmukh, Atul S
Mann, Matthias
Jensen, Lars Juhl
Meissner, Felix
Cox, Juergen
AuthorAffiliation Max-Planck-Institute of Biochemistry
The Novo Nordisk Foundation Center for Protein Research, Faculty of Health and Medical Sciences
University of Copenhagen
Department of Proteomics and Signal Transduction
AuthorAffiliation_xml – name: Max-Planck-Institute of Biochemistry
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  givenname: Lars Juhl
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Issue 11
Keywords insulin signaling
glucose uptake
mass spectrometry
myokines
metabolism
secretome
diabetes
palmitic acid
quantitative proteomics
obesity
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Snippet Skeletal muscle has emerged as an important secretory organ that produces so-called myokines, regulating energy metabolism via autocrine, paracrine, and...
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SubjectTerms Amino Acid Sequence
Animals
Cell Differentiation
Cell Line
Computational Biology - methods
Cytokines - genetics
Cytokines - isolation & purification
Cytokines - secretion
Gene Expression Regulation
Glucose - metabolism
Glucose - pharmacology
Insulin - metabolism
Insulin - pharmacology
Insulin Resistance
Insulin-Like Growth Factor Binding Proteins - genetics
Insulin-Like Growth Factor Binding Proteins - isolation & purification
Insulin-Like Growth Factor Binding Proteins - secretion
Intercellular Signaling Peptides and Proteins - genetics
Intercellular Signaling Peptides and Proteins - isolation & purification
Intercellular Signaling Peptides and Proteins - secretion
Metalloproteases - genetics
Metalloproteases - isolation & purification
Metalloproteases - secretion
Mice
Molecular Sequence Data
Muscle Fibers, Skeletal - cytology
Muscle Fibers, Skeletal - drug effects
Muscle Fibers, Skeletal - secretion
Myoblasts - cytology
Myoblasts - drug effects
Myoblasts - secretion
Palmitic Acid - pharmacology
Protein Sorting Signals - genetics
Protein Structure, Tertiary
Serum Albumin, Bovine - chemistry
Title Secretome Analysis of Lipid-Induced Insulin Resistance in Skeletal Muscle Cells by a Combined Experimental and Bioinformatics Workflow
URI http://dx.doi.org/10.1021/acs.jproteome.5b00720
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