Title Rad 9 modulates the P 21 WAF 1 pathway by direct association with p 53

Background: Previous studies suggest that human RAD9 (hRad9), encoding a DNA damage checkpoint molecule, which is frequently amplified in epithelial tumor cells of breast, lung, head and neck cancer, participates in regulation of the tumor suppressor p53-dependent transactivation of pro-survival P21WAF1. This study examined the exact mechanism of the hRad9 function, especially through the phosphorylation of the C-terminus, in the transcription regulation of P21WAF1. Results: The transfection of phosphorylation-defective hRAD9 mutants of C-terminus resulted in reduction of the p53-dependent P21WAF1 transactivation; the knockdown of total hRad9 elicited an increased P21WAF1 mRNA expression. Immunoprecipitation and a ChIP assay showed that hRad9 and p53 formed a complex and both were associated with two p53-consensus DNA-binding sequences in the 5' region of P21WAF1 gene. The association was reduced in the experiment of phosphorylation-defective hRAD9 mutants. Conclusion: The present study indicates the direct involvement of hRad9 in the p53-dependent P21WAF1 transcriptional mechanism, presumably via the phosphorylation sites, and alterations of the hRad9 pathway might therefore contribute to the perturbation of checkpoint activation in cancer cells. Background DNA damage checkpoints are signal transduction pathways that maintain the proper order of cell cycle events. When DNA is damaged or perturbed during replication, the cells respond by the activation of evolutionarily conserved signal transduction pathways that delay the progression of the cell cycle and induce repair of the damaged DNA. These signal transduction pathways include protein sensors that recognize aberrant DNA structures and activate kinases, thereby inducing phosphorylation cascades that ultimately lead to cell cycle arrest and DNA repair [1,2]. Failure of this cell cycle surveillance mechanism can Published: 21 May 2007 BMC Molecular Biology 2007, 8:37 doi:10.1186/1471-2199-8-37 Received: 2 February 2007 Accepted: 21 May 2007 This article is available from: http://www.biomedcentral.com/1471-2199/8/37 © 2007 Ishikawa et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. BMC Molecular Biology 2007, 8:37 http://www.biomedcentral.com/1471-2199/8/37