Ruolo funzionale della ciclina D3 nel differenziamento miogenico
Author(s)
Mezzaroma, Eleonora
Date Issued
February 20, 2009
Type
Doctoral Thesis
Abstract
The differentiation of skeletal muscle cells is characterized by a coordinated sequence of events that include irreversible exit from the cell cycle, the timely, ordered activation of muscle-specific gene expression, and the fusion of post mitotic myoblasts into multinucleated myotubes. This process is regulated by the basic helix-loop-helix family of myogenic factors (MRFs), which include MyoD, Myf5, myogenin and MRF4. Mitogenic signaling antagonizes by multiple mechanisms the activity of MRFs, which results in the absence of differentiation-specific gene expression in proliferating myoblasts. Conversely, the irreversible withdrawal of myoblasts from the cell cycle promotes their commitment into differentiation.
In earlier studies our laboratory has demonstrated that MyoD couples the onset of differentiation with cell-cycle arrest by inducing the expression of critical cell-cycle regulators, including retinoblastoma, the cyclin-dependent kinase inhibitor p21 and cyclin D3. In contrast to retinoblastoma protein (pRb) and p21, little is known about the function of cyclin D3 in the differentiation process. Our laboratory has previously shown that cyclin D3 is highly expressed and nearly totally associated with hypophosphorylated pRb in differentiated myotubes, whereas Rb-/- myocytes fail to accumulate the cyclin D3 protein, suggesting that pRb promotes cyclin D3 stabilization in differentiating myoblasts. In my study I have shown that, in proliferating C2 myoblasts, cyclin D3 displays rapid turnover, which is positively regulated trough glycogen synthase kinase 3β (GSK-3β)-mediated phosphorylation of cyclin D3 on Thr-283. The pRb-cyclin D3 complex formation in terminally differentiated myotubes hinders the access of GSK-3β to cyclin D3, thus inhibiting Thr-283 phosphorylation and the ensuing degradation of cyclin D3. My results indicate that the ectopic expression of a stabilized cyclin D3(T283A) mutant in C2 myoblasts increases the number of cells progressing toward differentiation by temporarily delaying muscle-specific gene expression, being anyway compatible with their terminal differentiation. Conversely, the expression of a stabilized double mutant cyclin D3-mLXCXE(dl250-272), unable to bind pRb, causes reduced proliferation and precocious differentiation of C2 myoblasts. Furthermore, cyclin D3-null embryonic fibroblasts display impaired MyoD-induced myogenic differentiation.
These results indicate that the pRb-dependent accumulation of cyclin D3 is functionally relevant to the process of skeletal muscle cell differentiation.
Additional information
Dottorato di ricerca in Genetica e biologia cellulare
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