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The dihydrofolate reductase (dhfr) promoter contains cis-acting element for the transcription factors Sp1 and E2F. Transcription of dhfr gene shows maximal activity during the G1/S phase of cell cycle. The member of the Sp1 transcriptional factor family can act as both negative and positive regulators of gene expression. There was a report that Sp1-Rb and E2F4-p130 complexes cooperate to establish stable repression of dhfr gene expression in CHOC400 cells. Here, we examined the role of HDAC in dhfr, cyclin E, and cyclin A gene regulation using the histone deacetylation inhibitor, trichostatin A (TSA) in U2OS and C33A cells, a Rb-positive human osteosarcoma cell line, and a Rb-negative cervical carcinoma cell line, respectively. When the dhfr promoter constructs were applied in U2OS cells, TSA markedly stimulated over 14-fold of dhfr promoter activity through dhfr-Sp1 sites by the deletion of an E2F element. In contrast, the deletion of dhfr-Sp1 binding sites completely abolished promoter stimulation by TSA. The dhfr promoter activity including dhfr-Sp1 sites increased only 2-fold in C33A cells. Promoter activity containing only dhfr-E2F site did not have much effect by the treatment of TSA in both U2OS and C33A cells. On the other hand, treatment with TSA induced significantly mRNA expression of dhfr and cyclin E, whereas levels of cyclin A decreased in U2OS cells, but had no effect in C33A cells. These results indicate that TSA have contradictory effect, activation of dhfr and cyclin E genes on G1 phase, and down-regulation of cyclin A on G2 phase through transcriptional regulation in U2OS cells.
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