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Specific knockdown of δ-sarcoglycan gene in C2C12 in vitro causes post-translational loss of other sarcoglycans without mechanical stress

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Abstract

The precise role of δ-sarcoglycan (SG) that is constitutively expressed in skeletal muscle cells and may serve for maintaining the sarcolemmal integrity has not been identified. The δ-SG protein is at first among SG complex. To specifically identify the role in C2C12 cells during the myogenesis, we screened several RNA interference (RNAi) candidates at first, and knocked down both levels of the mRNA and protein, employing adenovirus-mediated RNAi. We found no morphological alteration at both myoblast and myotube stages by suppression of δ-SG. The specific knockdown of δ-SG accompanied a concomitant decrease of α-, β-, and γ-SGs preserving normal levels of each transcript. As for the localization, α-, β-, and γ-SGs were weakly stained on the cell membrane in δ-SG knockdown cells, whereas each SG in control cell was localized both on the cell membrane and myoplasm abundantly. This enhanced post-translational loss would represent similitude of the progression of cardiomuscular diseases in vitro. Different from cardiac muscle cells, skeletal muscle cell culture without muscle contraction may imply that mechanical stress per se is not primarily involved in the progression of limb-girdle muscular dystrophy. Furthermore, we have observed translocation of calpain-2 to cell membrane in δ-SG knockdown cells, suggesting that Ca2+-sensitive proteases, calpains closely take part in post-translational proteolysis.

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Authors and Affiliations

  1. Department of Materials and Life Science, Faculty of Science and Engineering, Sophia University, Tokyo, 102-8554, Japan

    Michiyo Honda, Mari Hosoda, Nobuyuki Kanzawa & Takahide Tsuchiya

  2. Department of Organ Pathophysiology and Internal Medicine, University of Tokyo, Tokyo, 113-0033, Japan

    Teruhiko Toyo-oka

  3. Department of Molecular Cardiology, Division of Biofunctional Sciences, Tohoku University Bioengineering Organization (TUBERO), Sendai, 980-8575, Japan

    Teruhiko Toyo-oka

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  1. Michiyo Honda
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  2. Mari Hosoda
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Correspondence to Michiyo Honda.

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Honda, M., Hosoda, M., Kanzawa, N. et al. Specific knockdown of δ-sarcoglycan gene in C2C12 in vitro causes post-translational loss of other sarcoglycans without mechanical stress. Mol Cell Biochem 323, 149–159 (2009). https://doi.org/10.1007/s11010-008-9975-2

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