Department of Medicine and Experimental Oncology, University of Turin, Turin, Italy.
J Cell Mol Med. 2010 Jun;14(6A):1347-57. doi: 10.1111/j.1582-4934.2009.00966.x. Epub 2009 Nov 13.
In human cells the length of telomeres depends on telomerase activity. This activity and the expression of the catalytic subunit of human telomerase reverse transcriptase (hTERT) is strongly up-regulated in most human cancers. hTERT expression is regulated by different transcription factors, such as c-Myc, Mad1 and Sp1. In this study, we demonstrated that 15d-PG J2 and rosiglitazone (an endogenous and synthetic peroxisome proliferators activated receptor gamma (PPARgamma) ligand, respectively) inhibited hTERT expression and telomerase activity in CaCo-2 colon cancer cells. Moreover, both ligands inhibited c-Myc protein expression and its E-box DNA binding activity. Additionally, Mad1 protein expression and its E-box DNA binding activity were strongly increased by 15d-PG J2 and, to a lesser extent, by rosiglitazone. Sp1 transcription factor expression and its GC-box DNA binding activity were not affected by both PPARgamma ligands. Results obtained by transient transfection of CaCo-2 cells with pmaxFP-Green-PRL plasmid constructs containing the functional hTERT core promoter (including one E-box and five GC-boxes) and its E-box deleted sequences, cloned upstream of the green fluorescent protein reporter gene, demonstrated that 15d-PG J2, and with minor effectiveness, rosiglitazone, strongly reduced hTERT core promoter activity. E-boxes for Myc/Mad/Max binding showed a higher activity than GC-boxes for Sp1. By using GW9662, an antagonist of PPARgamma, we demonstrated that the effects of 15d-PG J2 are completely PPARgamma independent, whereas the effects of rosiglitazone on hTERT expression seem to be partially PPARgamma independent. The regulation of hTERT expression by 15d-PG J2 and rosiglitazone, through the modulation of the Myc/Max/Mad1 network, may represent a new mechanism of action of these substances in inhibiting cell proliferation.
在人类细胞中，端粒的长度取决于端粒酶的活性。这种活性和人端粒酶逆转录酶（hTERT）的催化亚基的表达在大多数人类癌症中被强烈上调。hTERT 的表达受到不同转录因子的调节，如 c-Myc、Mad1 和 Sp1。在这项研究中，我们证明 15d-PGJ2 和罗格列酮（分别是内源性和合成过氧化物酶体增殖物激活受体 γ（PPARγ）配体）抑制了 CaCo-2 结肠癌细胞中的 hTERT 表达和端粒酶活性。此外，两种配体均抑制了 c-Myc 蛋白表达及其 E 盒 DNA 结合活性。此外，15d-PGJ2 强烈增加了 Mad1 蛋白表达及其 E 盒 DNA 结合活性，而罗格列酮的作用则较弱。Sp1 转录因子表达及其 GC 盒 DNA 结合活性不受两种 PPARγ 配体的影响。用包含功能性 hTERT 核心启动子（包括一个 E 盒和五个 GC 盒）及其 E 盒缺失序列的 pmaxFP-Green-PRL 质粒构建体瞬时转染 CaCo-2 细胞后获得的结果表明，15d-PGJ2 强烈降低了 hTERT 核心启动子活性，而罗格列酮的作用则较弱。用于 Myc/Mad/Max 结合的 E 盒比用于 Sp1 的 GC 盒具有更高的活性。通过使用 GW9662，一种 PPARγ 拮抗剂，我们证明 15d-PGJ2 的作用完全不依赖于 PPARγ，而罗格列酮对 hTERT 表达的影响似乎部分不依赖于 PPARγ。15d-PGJ2 和罗格列酮通过调节 Myc/Max/Mad1 网络对 hTERT 表达的调节，可能代表了这些物质抑制细胞增殖的一种新的作用机制。
