Sp1 cooperates with c-Myc to activate transcription of the human telomerase reverse transcriptase gene (hTERT)

Abstract

Telomerase activation is thought to be a critical step in cellular immortalization and carcinogenesis. The human telomerase catalytic subunit (hTERT) is a rate limiting determinant of the enzymatic activity of human telomerase. In the previous study, we identified the proximal 181 bp core promoter responsible for transcriptional activity of the hTERT gene. To identify the regulatory factors of transcription, transient expression assays were performed using hTERT promoter reporter plasmids. Serial deletion assays of the core promoter revealed that the 5'-region containing the E-box, which binds Myc/Max, as well as the 3'-region containing the GC-box, which binds Sp1, are essential for transactivation. The mutations introduced in the E-box or GC-box significantly decreased transcriptional activity of the promoter. Overexpression of Myc/Max or Sp1 led to significant activation of transcription in a cell type-specific manner, while Mad/Max introduction repressed it. However, the effects of Myc/Max on transactivation were marginal when Sp1 sites were mutated. Western blot analysis using various cell lines revealed a positive correlation between c-Myc and Sp1 expression and transcriptional activity of hTERT. Using fibroblast lineages in different stages of transformation, we found that c-Myc and Sp1 were induced to a dramatic extent when cells overcame replicative senescence and obtained immortal characteristics, in association with telomerase activation. These findings suggest that c-Myc and Sp1 cooperatively function as the major determinants of hTERT expression, and that the switching functions of Myc/Max and Mad/Max might also play roles in telomerase regulation.

Infectivity enhanced, hTERT promoter-based conditionally replicative adenoviruses are useful for SCLC treatment

“…The mechanisms by which the hTERT core promoter works so differentially in tumors and normal cells are still unclear. However, it has been reported that c-Myc, 29,[53][54][55][56][57] Sp1 57 and estrogen 58,59 activated hTERT promoter, but p53, 60 WT-1 61 and Mad1 62 were important factors for the Figure 10 Ad5/3hTERTE1 suppressed tumor growth in vivo. Intact SBC5 cells (1 Â 10 7 ) were injected s.c. into nude mice.…”

Inhibition of telomerase activity in malignant glioma cells correlates with their sensitivity to temozolomide

“…On the other hand, it has been recently demonstrated that arsenic downregulates the promoter activity of hTERT by interfering with both c-Myc and Sp1 binding sites (Chou et al, 2001). Although Sp1 and c-Myc cooperatively activate hTERT expression (Kyo et al, 2000), it is possible that TMZ or E2 directly or indirectly suppresses the transactivation function of Sp1 itself.…”

“…Various lengths of DNA fragments upstream of the initiating ATG codon of the hTERT promoter gene were PCR amplified and inserted into luciferase reporter vector pGL3-Basic (Promega CORP-BTC, Madison, WI, USA). For the construction of reporter plasmids containing substitution mutations in factor binding sites of c-Myc or Sp1, site-specific mutagenesis was performed by a PCR-based protocol as described previously (Kyo et al, 2000). Cells were plated at a density of 5 Â 10 4 cells ml À1 day prior to TMZ treatment.…”

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].…”