Identification and characterization of negative regulatory elements of the human telomerase catalytic subunit (hTERT) gene promoter: possible role of MZF-2 in transcriptional repression of hTERT

Human telomerase reverse transcriptase (hTERT) is a catalytic subunit of human telomerase and is a critical determinant of the enzymatic activity of telomerase. Expression of hTERT is known to be regulated mainly at the transcriptional level. In the present study, using transient expression assays, we identified a 400 bp silencer of the hTERT promoter between -776 and -378 upstream of the proximal core promoter. The inhibitory effects of this silencer were enhanced with cellular differentiation. A computer-assisted homology search identified multiple binding motifs for myeloid-specific zinc finger protein 2 (MZF-2) within this region. Mutation introduced in these sites resulted in significant activation of hTERT transcription. Gel shift assays demonstrated that MZF-2 proteins specifically bound to these sites. Overexpression of MZF-2 in cells led to down-regulation of hTERT transcription as well as telomerase activity. These findings suggest that the 400 bp region upstream of the hTERT core promoter that we identified functions as a negative regulatory region and that MZF-2 may be an effector of negative regulation of hTERT.

[1]  S. Kyo,et al.  Human telomerase reverse transcriptase as a critical determinant of telomerase activity in normal and malignant endometrial tissues , 1999, International journal of cancer.

[2]  et al.,et al.  The RNA component of human telomerase , 1995, Science.

[3]  M. Yutsudo,et al.  Sp1 cooperates with c-Myc to activate transcription of the human telomerase reverse transcriptase gene (hTERT). , 2000, Nucleic acids research.

[4]  C. Harley,et al.  Extension of life-span by introduction of telomerase into normal human cells. , 1998, Science.

[5]  S. Nagata,et al.  Myeloid-specific transcriptional activation by murine myeloid zinc-finger protein 2. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[6]  S. Kyo,et al.  hTERT is a critical determinant of telomerase activity in renal‐cell carcinoma , 1998, International journal of cancer.

[7]  R. Weinberg,et al.  hEST2, the Putative Human Telomerase Catalytic Subunit Gene, Is Up-Regulated in Tumor Cells and during Immortalization , 1997, Cell.

[8]  S. Kyo,et al.  Expression of human telomerase subunits and correlation with telomerase activity in urothelial cancer. , 1998, Clinical cancer research : an official journal of the American Association for Cancer Research.

[9]  S. Kyo,et al.  Expression of human telomerase subunits and correlation with telomerase activity in cervical cancer. , 1998, Cancer research.

[10]  R. Dalla‐Favera,et al.  Direct activation of TERT transcription by c-MYC , 1999, Nature Genetics.

[11]  C. Greider.  Chromosome first aid , 1991, Cell.

[12]  Gregg B. Morin,et al.  Reconstitution of human telomerase with the template RNA component hTR and the catalytic protein subunit hTRT , 1997, Nature Genetics.

[13]  Expression of human telomerase subunits in ovarian malignant, borderline and benign tumors , 1999, International journal of cancer.

[14]  J. Feng,et al.  An altered repertoire of fos/jun (AP-1) at the onset of replicative senescence. , 1992, Experimental cell research.

[15]  J. Steitz,et al.  Telomere shortening associated with chromosome instability is arrested in immortal cells which express telomerase activity. , 1992, The EMBO journal.

[16]  P. Chomczyński,et al.  Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. , 1987, Analytical biochemistry.

[17]  A. Matsuura,et al.  TLP1: A Gene Encoding a Protein Component of Mammalian Telomerase Is a Novel Member of WD Repeats Family , 1997, Cell.

[18]  C. Harley,et al.  Telomerase activity in human ovarian carcinoma. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[19]  C. Greider,et al.  Telomere length regulation. , 1996, Annual review of biochemistry.

[20]  C B Harley,et al.  Telomerase catalytic subunit homologs from fission yeast and human. , 1997, Science.

[21]  F. Ishikawa,et al.  Telomerase activation by hTRT in human normal fibroblasts and hepatocellular carcinomas , 1998, Nature Genetics.

[22]  Thorsten Heinzel,et al.  Ligand-independent repression by the thyroid hormone receptor mediated by a nuclear receptor co-repressor , 1995, Nature.

[23]  M. Yutsudo,et al.  Cloning of human telomerase catalytic subunit (hTERT) gene promoter and identification of proximal core promoter sequences essential for transcriptional activation in immortalized and cancer cells. , 1999, Cancer research.

[24]  J. Yudkin.  Sugar and Disease , 1972, Nature.

[25]  C B Harley,et al.  Specific association of human telomerase activity with immortal cells and cancer. , 1994, Science.

[26]  Lea Harrington,et al.  A Mammalian Telomerase-Associated Protein , 1997, Science.

[27]  Y. Cong,et al.  The human telomerase catalytic subunit hTERT: organization of the gene and characterization of the promoter. , 1999, Human molecular genetics.