[Withdrawn] Molecular mechanisms behind skeletal muscle wasting
Date of Issue2012
School of Biological Sciences
Myostatin, a member of TGF-β superfamily, is a potent negative regulator of skeletal muscle growth and development. Current literature demonstrates that elevated levels of circulatory Myostatin activates the ubiquitin-proteasome pathway, which leads to skeletal muscle wasting. However, the exact signaling mechanism by which Myostatin regulates the activity of this proteolytic pathway in skeletal muscles is unclear. To characterize the role of Myostatin during skeletal muscle wasting, several lines of investigation were undertaken both in vitro and in vivo. Molecular analysis carried out in C2C12 myotubes and Smad3-null muscle indicated that Myostatin signals through Smad3 to activate the E3 ligase Atrogin-1, which in turn degrades sarcomeric proteins, such as Myosin heavy chain. In addition, Atrogin-1 also degrades myogenic differentiation factor (MyoD) and translation machinery components in response to Myostatin signaling. Since increased levels of Myostatin have been reported during cancer cachexia, we next analyzed if tumor cells are a source of Myostatin. Proteomic analysis of conditioned medium confirmed that Myostatin is indeed a tumoral factor secreted by not only C26 colon cancer cells but also by many human and murine neoplasms. Treating differentiated C2C12 myotubes with the C26 conditioned medium resulted in myotubular atrophy due to the up-regulation of muscle-specific E3 ligases Atrogin-1 and MuRF1 and enhanced activity of the ubiquitin-proteasome pathway. In addition to intracellular signaling pathways, we also analyzed the changes in the extracellular environment of myotubes exposed to procachectic insults. The nanoLC-MS/MS technique was employed to characterize the secreted proteins (secretome) of human primary myotubes treated with Myostatin. Proteomic analysis revealed that while the control myotubes secreted 332 proteins, treatment with Myostatin resulted in the secretion of 776 proteins. Further analysis of the secretome revealed increased secretion of Activin A, Thrombospondin-1, and Insulin Growth Factor Binding Protein 3 (IGF-BP3) in response to Myostatin, which may collectively aggravate skeletal muscle wasting.
DRNTU::Science::Biological sciences::Molecular biology