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Transient induction of telomerase expression mediates senescence and reduces tumorigenesis in primary fibroblasts.

ABSTRACT: Telomerase is an enzymatic ribonucleoprotein complex that acts as a reverse transcriptase in the elongation of telomeres. Telomerase activity is well documented in embryonic stem cells and the vast majority of tumor cells, but its role in somatic cells remains to be understood. Here, we report an unexpected function of telomerase during cellular senescence and tumorigenesis. We crossed Tert heterozygous knockout mice (mTert +/- ) for 26 generations, during which time there was progressive shortening of telomeres, and obtained primary skin fibroblasts from mTert +/+ and mTert -/- progeny of the 26th cross. As a consequence of insufficient telomerase activities in prior generations, both mTert +/+ and mTert -/- fibroblasts showed comparable and extremely short telomere length. However, mTert -/- cells approached cellular senescence faster and exhibited a significantly higher rate of malignant transformation than mTert +/+ cells. Furthermore, an evident up-regulation of telomerase reverse-transcriptase (TERT) expression was detected in mTert +/+ cells at the presenescence stage. Moreover, removal or down-regulation of TERT expression in mTert +/+ and human primary fibroblast cells via CRISPR/Cas9 or shRNA recapitulated mTert -/- phenotypes of accelerated senescence and transformation, and overexpression of TERT in mTert -/- cells rescued these phenotypes. Taking these data together, this study suggests that TERT has a previously underappreciated, protective role in buffering senescence stresses due to short, dysfunctional telomeres, and preventing malignant transformation.

SUBMITTER: Sun L

PROVIDER: S-EPMC6754593 | biostudies-literature | 2019 Sep

REPOSITORIES: biostudies-literature

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Transient induction of telomerase expression mediates senescence and reduces tumorigenesis in primary fibroblasts.

Sun Linlin L Chiang Jeffrey Y JY Choi Ji Young JY Xiong Zheng-Mei ZM Mao Xiaojing X Collins Francis S FS Hodes Richard J RJ Cao Kan K

Proceedings of the National Academy of Sciences of the United States of America 20190903 38

Telomerase is an enzymatic ribonucleoprotein complex that acts as a reverse transcriptase in the elongation of telomeres. Telomerase activity is well documented in embryonic stem cells and the vast majority of tumor cells, but its role in somatic cells remains to be understood. Here, we report an unexpected function of telomerase during cellular senescence and tumorigenesis. We crossed Tert heterozygous knockout mice (mTert+/- ) for 26 generations, during which ti ...[more]

PMID: 31481614

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Project description:Cyclin Dependent Kinase-2 Associated Protein-1 (CDK2AP1) is known to be a tumor suppressor that plays a role in cell cycle regulation by sequestering monomeric CDK2, and targeting it for proteolysis. A reduction of CDK2AP1 expression is considered to be a negative prognostic indicator in patients with oral squamous cell carcinoma and also associated with increased invasion in human gastric cancer tissue. CDK2AP1 overexpression was shown to inhibit growth, reduce invasion and increase apoptosis in prostate cancer cell lines. In this study, we investigated the effect of CDK2AP1 downregulation in primary human dermal fibroblasts. Using a short-hairpin RNA to reduce its expression, we found that knockdown of CDK2AP1 in primary human fibroblasts resulted in reduced proliferation and in the induction of senescence associated beta-galactosidase activity. CDK2AP1 knockdown also resulted in a significant reduction in the percentage of cells in the S phase and an accumulation of cells in the G1 phase of the cell cycle. Immunocytochemical analysis also revealed that the CDK2AP1 knockdown significantly increased the percentage of cells that exhibited ?-H2AX foci, which could indicate presence of DNA damage. CDK2AP1 knockdown also resulted in increased mRNA levels of p53, p21, BAX and PUMA and p53 protein levels. In primary human fibroblasts in which p53 and CDK2AP1 were simultaneously downregulated, there was: (a) no increase in senescence associated beta-galactosidase activity, (b) decrease in the number of cells in the G1-phase and increase in number of cells in the S-phase of the cell cycle, and (c) decrease in the mRNA levels of p21, BAX and PUMA when compared with CDK2AP1 knockdown only fibroblasts. Taken together, this suggests that the observed phenotype is p53 dependent. We also observed a prominent increase in the levels of ARF protein in the CDK2AP1 knockdown cells, which suggests a possible role of ARF in p53 stabilization following CDK2AP1 knockdown. Altogether, our results show that knockdown of CDK2AP1 in primary human fibroblasts reduced proliferation and induced premature senescence, with the observed phenotype being p53 dependent.

Dataset Information

Transient induction of telomerase expression mediates senescence and reduces tumorigenesis in primary fibroblasts.

Publications

Transient induction of telomerase expression mediates senescence and reduces tumorigenesis in primary fibroblasts.

Similar Datasets

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