Dynamics of the skeletal muscle secretome during myoblast differentiation

TY - JOUR

T1 - Dynamics of the skeletal muscle secretome during myoblast differentiation

AU - Henningsen, Jeanette

AU - Rigbolt, Kristoffer T G

AU - Blagoev, Blagoy

AU - Pedersen, Bente Klarlund

AU - Kratchmarova, Irina

PY - 2010/11/1

Y1 - 2010/11/1

N2 - During recent years, increased efforts have focused on elucidating the secretory function of skeletal muscle. Through secreted molecules, skeletal muscle affects local muscle biology in an auto/paracrine manner as well as having systemic effects on other tissues. Here we used a quantitative proteomics platform to investigate the factors secreted during the differentiation of murine C2C12 skeletal muscle cells. Using triple encoding stable isotope labeling by amino acids in cell culture, we compared the secretomes at three different time points of muscle differentiation and followed the dynamics of protein secretion. We identified and quantitatively analyzed 635 secreted proteins, including 35 growth factors, 40 cytokines, and 36 metallopeptidases. The extensive presence of these proteins that can act as potent signaling mediators to other cells and tissues strongly highlights the important role of the skeletal muscle as a prominent secretory organ. In addition to previously reported molecules, we identified many secreted proteins that have not previously been shown to be released from skeletal muscle cells nor shown to be differentially released during the process of myogenesis. We found 188 of these secreted proteins to be significantly regulated during the process of myogenesis. Comparative analyses of selected secreted proteins revealed little correlation between their mRNA and protein levels, indicating pronounced regulation by posttranscriptional mechanisms. Furthermore, analyses of the intracellular levels of members of the semaphorin family and their corresponding secretion dynamics demonstrated that the release of secreted proteins is tightly regulated by the secretory pathway, the stability of the protein, and/or the processing of secreted proteins. Finally, we provide 299 unique hydroxyproline sites mapping to 48 distinct secreted proteins and have discovered a novel hydroxyproline motif.

AB - During recent years, increased efforts have focused on elucidating the secretory function of skeletal muscle. Through secreted molecules, skeletal muscle affects local muscle biology in an auto/paracrine manner as well as having systemic effects on other tissues. Here we used a quantitative proteomics platform to investigate the factors secreted during the differentiation of murine C2C12 skeletal muscle cells. Using triple encoding stable isotope labeling by amino acids in cell culture, we compared the secretomes at three different time points of muscle differentiation and followed the dynamics of protein secretion. We identified and quantitatively analyzed 635 secreted proteins, including 35 growth factors, 40 cytokines, and 36 metallopeptidases. The extensive presence of these proteins that can act as potent signaling mediators to other cells and tissues strongly highlights the important role of the skeletal muscle as a prominent secretory organ. In addition to previously reported molecules, we identified many secreted proteins that have not previously been shown to be released from skeletal muscle cells nor shown to be differentially released during the process of myogenesis. We found 188 of these secreted proteins to be significantly regulated during the process of myogenesis. Comparative analyses of selected secreted proteins revealed little correlation between their mRNA and protein levels, indicating pronounced regulation by posttranscriptional mechanisms. Furthermore, analyses of the intracellular levels of members of the semaphorin family and their corresponding secretion dynamics demonstrated that the release of secreted proteins is tightly regulated by the secretory pathway, the stability of the protein, and/or the processing of secreted proteins. Finally, we provide 299 unique hydroxyproline sites mapping to 48 distinct secreted proteins and have discovered a novel hydroxyproline motif.

KW - Amino Acid Sequence

KW - Animals

KW - Cell Differentiation

KW - Cell Line

KW - Isotope Labeling

KW - Mass Spectrometry

KW - Mice

KW - Molecular Sequence Data

KW - Muscle Proteins

KW - Muscle, Skeletal

KW - Myoblasts

KW - Protein Isoforms

KW - Proteomics

KW - Semaphorins

U2 - 10.1074/mcp.M110.002113

DO - 10.1074/mcp.M110.002113

M3 - Journal article

C2 - 20631206

SN - 1535-9476

VL - 9

SP - 2482

EP - 2496

JO - Molecular and Cellular Proteomics

JF - Molecular and Cellular Proteomics

IS - 11

ER -