Transcriptional Regulation of Mitotic Checkpoint Gene MAD1 by p53*
- ‡ To whom correspondence should be addressed: Dept. of Biochemistry, the University of Hong Kong, 3rd Floor, Laboratory Block, Faculty of Medicine Bldg., 21 Sassoon Rd., Hong Kong, China. Tel.: 852-2819-9491; Fax: 852-2855-1254; E-mail: dyjin{at}hkucc.hku.hk.
Abstract
p53 regulates a number of genes through transcriptional activation and repression. p53-dependent mitotic checkpoint has been described, but the underlying mechanism is still obscure. Here we examined the effect of p53 on the expression of a human mitotic checkpoint protein, Mitosis Arrest Deficiency 1 (MAD1), in cultured human cells. The expression of MAD1 was reduced when the cells were overexpressing exogenously introduced wild-type p53. The same reduction was also observed when the cells were treated with anticancer agents 5-fluorouracil and cisplatin or were irradiated with UV. Consistently, MAD1 promoter activity diminished in a dose-dependent manner when induced by p53, indicating that p53 repressed MAD1 at a transcriptional level. Intriguingly, several tumor hot spot mutations in p53 (V143A, R175H, R248W, and R273H) did not abolish the ability of p53 to repress MAD1 expression. By serial truncation of the MAD1 promoter, we confined the p53-responsive element to a 38-bp region that represents a novel sequence distinct from the known p53 consensus binding site. Trichostatin A, a histone deacetylase inhibitor, relieved the p53 transrepression activity on MAD1. Chromatin immunoprecipitation assay revealed that p53, histone deacetylase 1, and co-repressor mSin3a associated with the MAD1 promoter in vivo. Taken together, our findings suggest a regulatory mechanism for the mitotic checkpoint in which MAD1 is inhibited by p53.
Footnotes
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↵1 The abbreviations used are: APC, anaphase-promoting complex; MAD, mitotic arrest deficiency; 5-FU, 5-fluorouracil; TSA, trichostatin A; HDAC, histone deacetylase; CMV, cytomegalovirus; CAT, chloramphenicol acetyltransferase; ChIP, chromatin immunoprecipitation; BUB, budding uninhibited by benomyl; MPS, monopolar spindle; wt, wild type; SAGE, serial analysis of gene expression.
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↵* This work was supported by National Institutes of Health Research Grant D43 TW06186 (to D.-Y. J.) funded by the Fogarty International Center. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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- Received July 5, 2003.
- The American Society for Biochemistry and Molecular Biology, Inc.
