MyoD-Induced Expression of p21 Inhibits Cyclin-Dependent Kinase Activity upon Myocyte Terminal Differentiation

Abstract

The terminal differentiation of C2C12 skeletal muscle cells involves the activation of unique sets of genes and an irreversible withdrawal from the cell cycle. This process is associated with a decrease in cdk2 activity in cell extracts. The decrease in cdk2 activity correlates with diminished levels of cdk2 and cyclin A and with a marked induction of the p21 cyclin-dependent kinase (cdk) inhibitor. The upregulation of p21 occurred at the levels of mRNA and protein, and p21 formed a complex with the cyclin kinases in myotubes. Further, the immunodepletion of p21 from myotube extracts neutralized the heat-stable cdk2 inhibitory activity that was induced upon myogenic differentiation. The levels of p21 mRNA, protein, and activity remained constant in myotubes when they were reexposed to mitogen-rich growth medium, indicating that permanent changes in the cell's genetic program contribute to its sustained expression following terminal differentiation. Indeed, 10T1/2 fibroblasts transformed with the myogenic factor MyoD, but not the parental multipotent cells, upregulated p21 transcript levels when induced to differentiate by serum withdrawal, demonstrating that the upregulation is an integral feature of myogenic commitment and differentiation. The functional consequences of this upregulation were indicated by ectopically expressing p21 in myoblasts; this was sufficient for cell cycle arrest in mitogen-rich growth medium. The induction and sustained expression of p21 appears to be a contributory mechanism by which myocytes irreversibly exit the cell cycle upon terminal differentiation.

p21WAF1 induces permanent growth arrest and enhances differentiation, but does not alter apoptosis in PC12 cells

“…In examining the process of cell-cycle withdrawal accompanying differentiation, p21 levels have been shown to increase during NGF treatment of neuroblastoma (Poluha et al, 1996) and PC12 cells (Yan and Zi , 1995;van Grunsven et al, 1996a). This correlation is not limited to neural cells, as induction of p21 mRNA and protein has been detected during myogenic (Guo et al, 1995;Halevy et al, 1995), keratinocytic (Missero et al, 1995), promyelocytic (HL-60) (Jiang et al, 1994;Steinman et al, 1994), and human melanoma cell di erentiation (Jiang et al, 1995).…”

Rhabdomyosarcoma – working out the pathways

“…Deletion of p21 was not able to substitute for p53 de®ciency in the rescue of the MDM2 null embryonic lethality, described above (Montes de Oca Luna et al, 1997). Moreover, although the p21 CIP1 gene is transcriptionally activated by p53 (el-Deiry et al, 1993), p21 CIP1 is also upregulated during myogenic di erentiation by a p53-independent mechanism thought to involve MyoDmediated transactivation (Parker et al, 1995;Guo et al, 1995;Halevy et al, 1995;Missero et al, 1995). It has also been shown that p21 CIP1 is poorly expressed in RMS cells lines, concomitant with both MyoD and p53 inactivity, and failure to G 1 arrest (Otten et al, 1997).…”

Genome‐wide examination of myoblast cell cycle withdrawal during differentiation

“…MyoD can promote cell cycle arrest by induction of the cyclin-dependent kinase inhibitor, p21 (Guo et al, 1995;Halevy et al, 1995), which along with other Cip1/Kip1 family members, p57 and p27, inhibits a wide range of cyclin-dependent kinases (CDKs) essential for cell cycle progression (Sherr and Roberts, 1999). It is generally accepted that induction of p21 follows myogenin expression during myoblast differentiation and that a high level of p21 is required for cells to maintain the postmitotic state.…”