Association of hsp90 to the hTERT promoter is necessary for hTERT expression in human oral cancer cells
Association of hsp90 to the hTERT promoter is necessary for hTERT expression in human oral cancer cells
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DOI：
10.1093/carcin/bgn225
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发表时间：
2008-12-01
期刊：
影响因子：
4.7
通讯作者：
Kang, Mo K.
中科院分区：
文献类型：
--
作者：
Kim, Reuben H.;Kim, Roy;Kang, Mo K.
人端粒酶逆转录酶（human telomerase reverse transcriptase，hTERT）的表达增强是细胞永生化过程中经常发生的现象。本研究旨在探讨人口腔角质形成细胞永生化过程中hTERT启动子活性的调控机制。用Bmi-1和人乳头瘤病毒16型E6癌蛋白永生化正常人口腔角质形成细胞（NHOKs），建立端粒酶阳性的HOK-Bmi-1/E6细胞系。利用DNA-蛋白结合实验，我们发现热休克蛋白90（hsp 90）在体外与hTERT启动子发生物理相互作用。与衰老的NHOK相比，在端粒酶阳性的HOK-Bmi-1/E6细胞中检测到更强烈的hsp 90与启动子的相互作用。染色质免疫沉淀证实了在SCC 4细胞，端粒酶阳性的口腔癌细胞系，但不是在NHOK中的hsp 90和hTERT启动子之间的体内相互作用。为了确定这种相互作用的生理意义，SCC 4细胞暴露于格尔德霉素（GA），热休克蛋白90的竞争性抑制剂。GA暴露导致SCC 4细胞中端粒酶活性、hTERT启动子活性和hTERT信使RNA表达降低，即使在没有从头蛋白合成的情况下也是如此。此外，它消除了hTERT启动子区与hsp 90的体内相互作用，但不与Sp1或c-Myc。这些结果表明hsp 90和hTERT启动子之间的物理相互作用发生在端粒酶阳性细胞中，而不是在正常的人类细胞中，是必要的增强hTERT表达和端粒酶活性在癌细胞中。
Enhanced expression of human telomerase reverse transcriptase (hTERT) occurs frequently during cellular immortalization. The current study was undertaken to determine the mechanism regulating the hTERT promoter activity during cellular immortalization of human oral keratinocytes. Normal human oral keratinocytes (NHOKs) were immortalized with Bmi-1 and the E6 oncoprotein of human papillomavirus type 16 to establish the telomerase-positive HOK-Bmi-1/E6 cell line. Using DNA-protein-binding assay, we found that heat shock protein 90 (hsp90) physically interacts with the hTERT promoter in vitro. The hsp90 interaction with the promoter was detected more strongly in the telomerase-positive HOK-Bmi-1/E6 cells compared with that in senescing NHOK. Chromatin immunoprecipitation confirmed the in vivo interaction between hsp90 and the hTERT promoter in SCC4 cells, a telomerase-positive oral cancer cell line, but not in the NHOK. To determine the physiological significance of this interaction, SCC4 cells were exposed to geldanamycin (GA), a competitive inhibitor of hsp90. GA exposure led to decrease in telomerase activity, hTERT promoter activity and hTERT messenger RNA expression in SCC4 cells, even in the absence of de novo protein synthesis. Also, it abolished the in vivo interaction of the hTERT promoter region with hsp90 but not with Sp1 or c-Myc. These results indicate that physical interaction between hsp90 and the hTERT promoter occurs in telomerase-positive cells but not in normal human cells and is necessary for the enhanced hTERT expression and telomerase activity in cancer cells.