Deacetylation of the retinoblastoma tumour suppressor protein by SIRT1.
The activity of Rb (retinoblastoma protein) is regulated by phosphorylation and acetylation events. Active Rb is hypophosphorylated and acetylated on multiple residues. Inactivation of Rb involves concerted hyper-phosphorylation by cyclin-CDK (cyclin-dependent kinase) complexes combined with deacetylation of appropriate lysine residues within Rb. In the present study, using in vivo co-immunoprecipitation experiments, we identified mammalian SIRT1 (sirtuin 1) as a binding partner for Rb and its family members p107 and p130. Formation of Rb-SIRT1 complexes required the pocket domain of Rb. p300 catalysed the acetylation of Rb, and SIRT1 was a potent deacetylase for Rb. The ability of SIRT1 to catalyse the deacetylation of Rb was dependent on NAD and was inhibited by the SIRT1 inhibitor nicotinamide. Deacetylated lysine residues within Rb formed a domain similar to the SIRT1-targeted domain of the p53 tumour suppressor protein. Cultures of arrested cells, via contact inhibition or DNA damage, exhibited decreased Rb phosphorylation and increased Rb acetylation. Overexpression of SIRT1 in either confluent or etoposide-treated cells resulted in a significant reduction in Rb acetylation, which was restored with nicotinamide. Gene knockdown of SIRT1 by siRNA (short interfering RNA) produced an accumulation of acetylated Rb. This increase was augmented further when siRNA against SIRT1 was used in conjunction with nicotinamide. In conclusion, our results demonstrate that SIRT1 is an in vitro and in vivo deacetylase for the Rb tumour suppressor protein.
Wong
Sharon
S
Division of Molecular Oncology, Department of Internal Medicine, Siteman Cancer Center, Washington University School of Medicine, 660 South Euclid Ave, St Louis, MI 63110, USA.
Weber
Jason D
JD
D000107:Acetylation; D000595:Amino Acid Sequence; D000818:Animals; D045744:Cell Line, Tumor; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D005810:Multigene Family; D034622:RNA Interference; D016160:Retinoblastoma Protein / Q000378:metabolism*; D050720:Retinoblastoma-Like Protein p107 / Q000378:metabolism; D050717:Retinoblastoma-Like Protein p130 / Q000378:metabolism; D056564:Sirtuin 1; D037761:Sirtuins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*
C493838:Rbl2 protein, mouse; D016160:Retinoblastoma Protein; D050720:Retinoblastoma-Like Protein p107; D050717:Retinoblastoma-Like Protein p130; C470545:Sirt1 protein, mouse; D056564:Sirtuin 1; D037761:Sirtuins