Binding of the retinoblastoma protein is not the determinant for stable repression of some E2F-regulated promoters in muscle cells.
Permanent silencing of E2F-dependent genes is a hallmark of the irreversible cell cycle exit that characterizes terminally differentiated and senescent cells. The determinant of this silencing during senescence has been proposed to be the binding of the retinoblastoma protein Rb and the consequent methylation of H3K9. During ex vivo skeletal muscle differentiation, while most cells terminally differentiate and form myotubes, a subset of myoblasts remains quiescent and can be reinduced by growth factor stimulation to enter the cell cycle. Thus, differentiating cells are composed of two different populations: one in which E2F-dependent genes are permanently repressed and the other not. We observed that, in a manner reminiscent to senescent cells, permanent silencing of the E2F-dependent cdc6, dhfr, and p107 promoters in myotubes was associated with a specific increase in H3K9 trimethylation. To investigate the role of Rb in this process, we developed a reliable method to detect Rb recruitment by chromatin immunoprecipitation. Surprisingly, we observed that Rb was recruited to these promoters more efficiently in quiescent cells than in myotubes. Thus, our data indicate that during muscle differentiation, permanent silencing and H3K9 trimethylation of some E2F-dependent genes are not directly specified by Rb binding, in contrast to what is proposed for senescence.
Vandromme
Marie
M
Laboratoire de Biologie Cellulaire et Moléculaire du Contrôle de la Prolifération, Centre National de la Recherche Scientifique, Université Paul Sabatier, Toulouse Cedex, France.
Chailleux
Catherine
C
Escaffit
Fabrice
F
Trouche
Didier
D
D000818:Animals; D002453:Cell Cycle; D002454:Cell Differentiation; D002460:Cell Line; D016922:Cellular Senescence; D002843:Chromatin / Q000378:metabolism; D050684:E2F Transcription Factors / Q000378:metabolism*; D005784:Gene Amplification; D051379:Mice; D018482:Muscle, Skeletal / Q000166:cytology / Q000502:physiology*; D032446:Myoblasts / Q000166:cytology / Q000502:physiology*; D011401:Promoter Regions, Genetic; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D016160:Retinoblastoma Protein / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D002843:Chromatin; D050684:E2F Transcription Factors; D016160:Retinoblastoma Protein; D012313:RNA