Rb is required for progression through myogenic differentiation but not maintenance of terminal differentiation

Abstract

To investigate the requirement for pRb in myogenic differentiation, a floxed Rb allele was deleted either in proliferating myoblasts or after differentiation. Myf5-Cre mice, lacking pRb in myoblasts, died immediately at birth and exhibited high numbers of apoptotic nuclei and an almost complete absence of myofibers. In contrast, MCK-Cre mice, lacking pRb in differentiated fibers, were viable and exhibited a normal muscle phenotype and ability to regenerate. Induction of differentiation of Rb-deficient primary myoblasts resulted in high rates of apoptosis and a total inability to form multinucleated myotubes. Upon induction of differentiation, Rb-deficient myoblasts up-regulated myogenin, an immediate early marker of differentiation, but failed to down-regulate Pax7 and exhibited growth in low serum conditions. Primary myoblasts in which Rb was deleted after expression of differentiated MCK-Cre formed normal multinucleated myotubes that did not enter S-phase in response to serum stimulation. Therefore, Rb plays a crucial role in the switch from proliferation to differentiation rather than maintenance of the terminally differentiated state.

Looking proximally and distally: 100 years of limb regeneration and beyond

“…However, myonuclei of normal mammalian myotubes do not re-enter the cell cycle in response to serum stimulation, suggesting that pRb inactivation is not sufficient to reverse cell cycle suppression. This was confirmed by experiments in which pRb was deleted (Huh et al, 2004) or knocked down by si RNA (Pajcini et al, 2010) in myotubes derived from primary myoblasts. The nuclei of these myotubes failed to re-enter the cell cycle in response to serum stimulation.…”

“…Reversal of differentiation by serum stimulation was also reported for myotubes differentiated from CC42 muscle cell lines derived from embryonic lethal Rb À/À mice (Schneider et al, 1994). The differentiation of wild-type skeletal muscle requires pRb (Huh et al, 2004), but p107 can substitute for pRb in pRb À/À muscle differentiation (Schneider et al, 1994). This alternate differentiation pathway apparently does not involve placing a lock on cell cycle re-entry like it does in the differentiation of normal muscle.…”

Lysine methylation regulates the pRb tumour suppressor protein

“…One of the properties of pRb that influences differentiation relates to the need to exit the cell cycle before differentiation can occur, and the incompatibility between proliferation and differentiation (Chen and Wang, 2000;Classon and Harlow, 2002;Huh et al, 2004;van den Heuvel and Dyson, 2008). Thus, derivatives of pRb that exhibit compromised cell cycle arrest activity, similar to K873A, might be expected to possess defective differentiation-inducing activity, reflecting incomplete cell cycle exit rather than a deficit in a differentiation activity per se.…”

“…For example, Rb À/À mice exhibit embryonic lethality resulting from cell autonomous and non-cell autonomous effects, causing a lack of cellular differentiation and increased levels of apoptosis in diverse tissues (Clarke et al, 1992;Jacks et al, 1992;Lee et al, 1992;van den Heuvel and Dyson, 2008). in vitro, pRb drives differentiation in different lineages, including osteoblasts and myoblasts, which involves pRb-dependent repression and cell cycle exit, together with transcriptional activation of differentiation-inducing transcription factors, such as MyoD and CBFA1 (Gu et al, 1993;Thorburn et al, 1993;Lipinski and Jacks, 1999;Novitch et al, 1999;Thomas et al, 2001;Huh et al, 2004).…”

Role of p53 and Rb in Ovarian Cancer

“…In addition to cell cycle effects through E2F, RB also has wide-ranging and frequently poorly understood functions in several cellular processes, including control of cell death and differentiation and histone modification. For example, RB plays a role in the transition of proliferating myoblasts to differentiating myocytes (Huh et al, 2004) and differentiation of fetal liver macrophages by opposing inhibitory functions of Id2 on transcription factor PU.1 (Iavarone et al, 2004). Furthermore, inactivation of Rb results in p53-independent apoptotic death in the developing nervous system of the mouse (Macleod et al, 1996).…”