Suppression of cyclin D1 by hypoxia-inducible factor-1 via direct mechanism inhibits the proliferation and 5-fluorouracil-induced apoptosis of A549 cells.

Hypoxia-inducible factor (HIF) and cyclin D1 are both key mediators of cell growth and proliferation in normal and cancer cells. However, the interrelation between HIF and cyclin D1 remains unclear. In the present study, we observed the inverse correlation between cyclin D1 and HIF-1 in hypoxia condition. Overexpression of the dominant negative mutant of HIF-1alpha (DN-HIF) significantly enhanced cyclin D1 expression upon hypoxia or arsenite exposure, suggesting the negative regulation of cyclin D1 by HIF-1. Furthermore, we found that the impairment of HIF-1 increased cyclin D1 expression in A549 pulmonary cancer cells, which in turn promoted G1-S cell cycle transition and cell proliferation. Cyclin D1 expression was increased in s.c. xenograft of DN-HIF stably transfected A549 cells in nude mice compared with that of control cells. Chromatin immunoprecipitation assay revealed that HIF-1 was able to directly bind to the promoter region of cyclin D1, which indicates that the negative regulation of cyclin D1 by HIF-1 is through a direct mechanism. Inhibition of histone deacetylase (HDAC) by pretreatment of cells with trichostatin A or specific knockdown of HDAC7 by its shRNA antagonized the suppression of cyclin D1 by HIF-1, suggesting that HDAC7 is required for HIF-1-mediated cyclin D1 downregulation. Moreover, we found that 5-fluorouracil-triggered apoptosis of DN-HIF-transfected A549 cells was reduced by sicyclin D1 (cyclin D1-specific interference RNA) introduction, suggesting that clinical observation of HIF-1 overexpression-associated chemoresistance might be, at least partially, due to the negative regulation of cyclin D1.