Tumor-specific Activation of Human Telomerase Reverses Transcriptase Promoter Activity by Activating Enhancer-binding Protein-2β in Human Lung Cancer Cells

Abstract

The up-regulated expression and telomerase activity of human telomerase reverse transcriptase (hTERT) are hallmarks of tumorigenesis. The hTERT promoter has been shown to promote hTERT gene expression selectively in tumor cells but not in normal cells. However, little is known about how tumor cells differentially activate hTERT transcription and induce telomerase activity. In this study, we identified activating enhancerbinding protein-2␤ (AP-2␤) as a novel transcription factor that specifically binds to and activates the hTERT promoter in human lung cancer cells. AP-2␤ was detected in hTERT promoter DNA-protein complexes formed in nuclear extracts prepared only from lung cancer cells but not from normal cells. We verified the tumor-specific binding activity of AP-2␤ for the hTERT promoter in vitro and in vivo and detected high expression levels of AP-2␤ in lung cancer cells. We found that ectopic expression of AP-2␤ reactivated hTERT promoter-driven reporter green fluorescent protein (GFP) gene and endogenous hTERT gene expression in normal cells, enhanced GFP gene expression in lung cancer cells, and prolonged the life span of primary lung bronchial epithelial cells. Furthermore, we found that inhibition of endogenous AP-2␤ expression by AP-2␤ gene-specific small interfering RNAs effectively attenuated hTERT promoter-driven GFP expression, suppressed telomerase activity, accelerated telomere shortening, and inhibited tumor cell growth by induction of apoptosis in lung cancer cells. Our results demonstrate the tumor-specific activation of the hTERT promoter by AP-2␤ and imply the potential of AP-2␤ as a novel tumor marker or a cancer therapeutic target.Telomerase is an RNA-dependent DNA polymerase ribonucleoprotein enzyme that synthesizes telomeres after cell division and maintains chromosomal stability, leading to cellular immortalization (1, 2). It comprises a template-containing RNA component and a human telomerase reverse transcriptase (hTERT) 2 in humans (3-6). The high activity of telomerase has been implicated in cellular immortalization, transformation, and oncogenesis. Most normal somatic cells do not display telomerase activity, whereas a high level of telomerase activity is detected in germinal cells, immortalized cell lines, and 85-90% of human cancer specimens (7-13). hTERT is the catalytic subunit of telomerase (4, 5). The elevated expression of hTERT is necessary to transform normal human cells into cancer cells and is regarded as a hallmark of tumorigenesis (14). The hTERT promoter has been showed to be capable of promoting both constitutive hTERT gene and transgene expression selectively in tumors but not in normal cells (15)(16)(17). This is largely attributed to the unique ability of tumor cells to up-regulate hTERT transcription. The expression of hTERT is tightly regulated by various cellular factors. For example, expression of c-Myc has been shown to induce the expression of hTERT, whereas the expression of SP1 (stimulating protein-1) suppresses it (18,19). Two E-boxes and an activating en...

Activator Protein-2β Promotes Tumor Growth and Predicts Poor Prognosis in Breast Cancer

“…Only one study based on cDNA microarray demonstrated the high expression of AP-2β in lobular breast tumor samples compared to ductal carcinomas [20]. Combined with the previous reports indicating AP-2β functionalized as oncogene to activate human telomerase reverses transcriptase and to up-regulate the transcription of the CRYAB gene through stabilizing p53 [21,22], most likely, we hypothesize the similarly tumorigenic role of AP-2β in breast cancer development, especially in advanced breast cancer.…”

“…However, to date, most studies focused on AP-2α and AP-2γ, and the reports about the other three family members, AP-2β, AP-2δ, and AP-2ε, were very rare. Our previous studies demonstrated the tumorigenic role of AP-2β in lung cancer by transcriptionally regulating human telomerase reverses transcriptase [22]. To expand this point, it might also play the same role in other cancer types, including breast cancer.…”

CDC5L Promotes hTERT Expression and Colorectal Tumor Growth

“…This observation is largely attributed to several key transcription factors in tumor cells that can up-regulate hTERT transcription [29]. To identify these potentially critical unknown factors, we successfully established a screening system that combined a streptavidin-agarose pulldown assay and high throughput proteomics [30]. One of the proteins identified using this systematic approach was CDC5L, which was expressed in hTERT promoter probe-protein complexes in the nuclear protein fractions prepared from colon cancer cells.…”

RuvBl2 cooperates with Ets2 to transcriptionally regulate hTERT in colon cancer

“…Similarly, a large number of TFs are known to activate hTERT transcription such as Sp1, c-Myc, GRHL2 and AP2 [16][17][18][19][20][21][22]. Of these, c-Myc has been directly observed to interact with E-boxes and its activation of hTERT transcription is partly dependent on chromatin configuration of the hTERT gene [23,24].…”