Acetylation of p53 at lysine 373/382 by the histone deacetylase inhibitor depsipeptide induces expression of p21(Waf1/Cip1).
Generally, histone deacetylase (HDAC) inhibitor-induced p21(Waf1/Cip1) expression is thought to be p53 independent. Here we found that an inhibitor of HDAC, depsipeptide (FR901228), but not trichostatin A (TSA), induces p21(Waf1/Cip1) expression through both p53 and Sp1/Sp3 pathways in A549 cells (which retain wild-type p53). This is demonstrated by measuring relative luciferase activities of p21 promoter constructs with p53 or Sp1 binding site mutagenesis and was further confirmed by transfection of wild-type p53 into H1299 cells (p53 null). That p53 was acetylated after depsipeptide treatment was tested by sequential immunoprecipitation/Western immunoblot analysis with anti-acetylated lysines and anti-p53 antibodies. The acetylated p53 has a longer half-life due to a significant decrease in p53 ubiquitination. Further study using site-specific antiacetyllysine antibodies and transfection of mutated p53 vectors (K319/K320/K321R mutated and K373R/K382R mutations) into H1299 cells revealed that depsipeptide specifically induces p53 acetylation at K373/K382, but not at K320. As assayed by coimmunoprecipitation, the K373/K382 acetylation is accompanied by a recruitment of p300, but neither CREB-binding protein (CBP) nor p300/CBP-associated factor (PCAF), to the p53 C terminus. Furthermore, activity associated with the binding of the acetylated p53 at K373/K382 to the p21 promoter as well as p21(Waf1/Cip1) expression is significantly increased after depsipeptide treatment, as tested by chromatin immunoprecipitations and Western blotting, respectively. In addition, p53 acetylation at K373/K382 is confirmed to be required for recruitment of p300 to the p21 promoter, and the depsipeptide-induced p53 acetylation at K373/K382 is unlikely to be dependent on p53 phosphorylation at Ser15, Ser20, and Ser392 sites. Our data suggest that p53 acetylation at K373/K382 plays an important role in depsipeptide-induced p21(Waf1/Cip1) expression.
Zhao
Ying
Y
Department of Biochemistry and Molecular Biology, Peking University Health Science Center, #38 Xueyuan Road, Beijing 100083, China.
Lu
Shaoli
S
Wu
Lipeng
L
Chai
Guolin
G
Wang
Haiying
H
Chen
Yingqi
Y
Sun
Jia
J
Yu
Yu
Y
Zhou
Wen
W
Zheng
Quanhui
Q
Wu
Mian
M
Otterson
Gregory A
GA
Zhu
Wei-Guo
WG
D000107:Acetylation / Q000187:drug effects; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics / Q000378:metabolism*; D047630:Depsipeptides / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D005786:Gene Expression Regulation / Q000187:drug effects*; D006207:Half-Life; D056572:Histone Deacetylase Inhibitors; D006801:Humans; D008239:Lysine / Q000378:metabolism*; D010766:Phosphorylation; D011401:Promoter Regions, Genetic / Q000235:genetics; D011499:Protein Processing, Post-Translational; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012694:Serine / Q000378:metabolism; D016329:Sp1 Transcription Factor / Q000378:metabolism; D013997:Time Factors; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism*; D050880:p300-CBP Transcription Factors / Q000378:metabolism
D050759:Cyclin-Dependent Kinase Inhibitor p21; D047630:Depsipeptides; D056572:Histone Deacetylase Inhibitors; D012333:RNA, Messenger; D016329:Sp1 Transcription Factor; D016159:Tumor Suppressor Protein p53; D012694:Serine; C087123:romidepsin; D050880:p300-CBP Transcription Factors; D008239:Lysine