Regulation of SV40 large T-antigen stability by reversible acetylation.
Reversible acetylation on protein lysine residues has been shown to regulate the function of both nuclear proteins such as histones and p53 and cytoplasmic proteins such as alpha-tubulin. To identify novel acetylated proteins, we purified several proteins by the affinity to an anti-acetylated-lysine antibody from cells treated with trichostatin A (TSA). Among the proteins identified, here we report acetylation of the SV40 large T antigen (T-Ag). The acetylation site was determined to be lysine-697, which is located adjacent to the C-terminal Cdc4 phospho-degron (CPD). Overexpression of the CBP acetyltransferase acetylated T-Ag, whereas HDAC1, HDAC3 and SIRT1 bound and deacetylated T-Ag. The acetylation and deacetylation occurred independently of p53, a binding partner of T-Ag, but the acetylation was enhanced in the presence of p53. T-Ag in the cells treated with TSA and NA or the acetylation mimic mutant (K697Q) became unstable in COS-7 cells, suggesting that acetylation regulates stability of T-Ag. Indeed, NIH3T3 cells stably expressing K697Q showed decreased anchorage-independent growth compared with those expressing wild type or the K697R mutant. These results demonstrate that acetylation destabilizes T-Ag and regulates the transforming activity of T-Ag in NIH3T3 cells.
Shimazu
T
T
Chemical Genetics Laboratory, RIKEN, Hirosawa 2-1, Wako, Saitama, Japan.
Komatsu
Y
Y
Nakayama
K I
KI
Fukazawa
H
H
Horinouchi
S
S
Yoshida
M
M
D000107:Acetylation; D000595:Amino Acid Sequence; D000818:Animals; D000952:Antigens, Polyomavirus Transforming / Q000737:chemistry / Q000378:metabolism*; D001483:Base Sequence; D019556:COS Cells; D002522:Chlorocebus aethiops; D002851:Chromatography, High Pressure Liquid; D017931:DNA Primers; D006655:Histone Deacetylases / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D056564:Sirtuin 1; D037761:Sirtuins / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization
D000952:Antigens, Polyomavirus Transforming; D017931:DNA Primers; C447939:SIRT1 protein, human; D056564:Sirtuin 1; D037761:Sirtuins; D006655:Histone Deacetylases