Abstract #3473: Down regulation of HDAC2 in senescent fibroblasts involves cellular senescence through induction of p53 independent p21cip1/sdi1 transactivation

Histone deacetylases (HDACs) and Histone deacetyltrasferases (HATs) are involved in regulating of chromatin structure and gene regulation. p21cip1/sdi1 is well know to up-regulated in human fibroblasts senescence. Treatment of Histone deacetylase inhibitor, Trichostatin A, dramatically induces p21 transcription and inhibits cell grwoth. However, the regulation of HDAC expression and their roles in cellular senescence are still unclear. Here we showed that HDAC2 expression was downregulated in senescent human fibroblasts at both mRNA and protein levels. Downregulation of HDAC2 expression in cellular senescence was restored by the ectopic expression of telomerase gene, hTERT. To monitoring HDAC2 promoter activity, we have cloned 4.1 kbp HDAC2 promoter and constructed reporter vector using pGL4 based vector. By using this vector for luciferase assay, HDAC2 was found to be transcriptionally downregulated in senescence. We also constructed p21 promoter reporter vector to monitoring p21 transactivation by green and red fluorescent proteins. We used three types of p21 promoters, 1.8kbp promoter containing two p53 binding sites, \#916; p53-p21promoter deleting one p53 binding site, double\#916;p53-p21promoter deleting both p53 binding sites. Using these reporter vectors, we constructed stable monitoring cells from TIG-3 fibroblasts. Transfection of three different siRNAs targeting HDAC2 into these p21 monitoring young fibroblast cells result in up-regulation of both green and red fluorescent proteins and inhibit cell growth. Interestingly, double\#916;p53-p21promoter monitoring cells also induce both green and red fluorescent proteins, indicating that p53 independent pathway involves in p21 transactivation in senescence. Promoter deletion analysis of HDAC2 promoter revealed that minimum 13 base region on promoter was involved in transcriptional activation of HDAC2, and transcription regulators that bind to this region was downregulated in senescent cells. We tried to identify the HDAC2 transcription activators by database search and gel shift assays, but known transcription factors did not bind to this region. These findings suggest that novel transcriptional regulators and telomere shortening are involved in HDAC2 transcription in senescent cells.

Citation Information: In: Proc Am Assoc Cancer Res; 2009 Apr 18-22; Denver, CO. Philadelphia (PA): AACR; 2009. Abstract nr 3473.