The Mechanism Of TERT Protecting The Chromosome Ends In Tumor Cells

Telomere is the natural structure of the chromosome ends, it consist of the repeat telomeric DNA sequence TTAGGG and the associated shelterin proteins complex. Telomere protects chromosomal ends and that has critical importance for genome stability and cell long-termed survival. Telomerase maintains the telomere structure through replicating and elongating the telomeric DNA. In humans, the regulatory mechanism of telomere maintenance by telomerase has been implicated in both ageing and cancer. In view of the significant roles of telomere and telomerase in cancer and cancer therapy, the 2009 Nobel Prize in Physiology or Medicine was awarded to the three American scientists who found the telomerase and clarification the structure of telomeres.Telomerase is a specialized RNA-protein complex with reverse transcriptase activity.The catalytic protein subunit (TERT) and the telomeric DNA template TR are the two core components of telomerase complex. TERT adds the repeat TTAGGG DNA sequences into chromosome ends, repairs the telomeric sequences which is lost due to the terminal replicate problem, therby ensuring the integrity of chromosomes. TERT gene expression is the rate-limiting step for telomerase activity in humans and is suppressed in almost all human normal somatic cells. Therefore, normal human cells lack telomerase activity, their telomeric DNA lengths decrease gradually with the increasing numbers of cell divisions, eventually telomeres shortening induce cellular replicative senescence. On the contrary, Depression of TERT expression occurs in almost all cancers and consequently reactivating telomerase to maintain the short cancerous telomeres and endowing cancer cells immortalization. Therefore, TERT has been identified as an important target for anti tumor therapy.In recent years, more and more studies have revealed that TERT has a variety of non-canonical function independent of its telomerase activity , such as anti apoptosis and tumor promotion. However, the molecular mechanisms underlying TERT to play these noval functions still remain largely unknown.In this study,we identified a noval action for TERT to play a critical role in tumor chromosomal ends protection through a way unrelated to is well-known telomerase enzymatical activity, thereby revealing a new pathway for cancer specific treatment based on TERT targeting. The major findings and significances of our studies are included as followings.1.TERT possesses a new function in chromosome ends protection in tumor cells Through the suppression of the TERT expression by the RNAi technology,we found that inhibition of TERT expression results in an acute DNA damage response activation at the ends of chromosomes in telomerase-positive tumor cells, while TERT inhibition does not affect the chromosome end protection mechanisms in TERT-immortalized normal human somatic cells, suggesting that such a new function for TERT is specifically present in cancerous cellular environment. Further study with a telomerase activity defect mutant DN-TERT indicated that the telomere protective function of TERT is independent of its telomerase activity, suggesting a new unknown function of TERT in the protection of abnormal chromosome ends in the cancer cells, which is important for the tumor cells survival.2.TERT protects tumor chromosomes ends by inhibiting the activation ATR-mediated single-stranded DNA damage response there.Through analyzing the roles of two main DNA damage monitors ATM and ATR in mediating the telomere dysfunction by TERT silencing, we found that inhibition of ATR significantly eliminated the acute activation of DNA damage signals in chromosome ends in tumor cells under the RNAi TERT inhibition.Therefore, these results suggest that TERT has a novel function in inhibiting ATR-mediated single-stranded DNA damage response in chromosome ends, and this is its main mechanism for chromosome ends protection. We further demonstrated that the tumor killing effect by TERT RNAi silencing is mediated through the ATR-ATM-Chk1-Chk2-P53 signaling pathways, thereby providing a key mechanisms for TERT to play a telomerase-unrelated anti apoptosis function in tumor cells.3.TERT plays the chromosomal end protective role in cancer cells through the TPP1 interaction.The finding that TERT silencing induces the activation of th ATR-dependent DNA single strand damage at telomeres suggests that TERT has a function in protecting the 3'telomeric overhang present at chromosomal ends. Pot1/TPP1 is known as main protective proteins for the 3'single-stranded telomeric overhang. It has been demonstrated that TPP1 could interact with TERT and is responsible for recruiting telomerase to telomeres in cancer cells. However, the biologic significance of TERT recruited by TPP1 in telomere maintenance is still poorly understood. We recently found that the interaction of Pinx-pot1 is also involved in telomerase telomere recruitment, and Pinx1 inhibition leads to telomere shorting in telomerase-positive cancer cells, but without inducing acute telomere dysfunction, suggesting that the PinX1-Pot1 interaction may account for directing TERT/telomerase to perform the telomere-length maintaning function on telomere ends. In this study, we further investigated the biologic significances of the TPP1-TERT interaction in telomere maintenance in human cancer cells. We provided direct evidences to demonstrate that the major function of the TPP1-TERT is to mediate TERT to exert the chromosomal ends protection in human cancer cells and such a mechanism is not required for chromosomal ends protection in normal human cells.4.The N-terminal(1-234aa) amino acids sequence represents a new functional domain for TERT to TPP1 interaction and protect telomeres in cancer cells.We have demonstrated that TERT have a novel role of protecting chromosome end independent of its known catalytic activity. Moreover, we identified the domain of this new function for TERT in cancer cells. TERT N-terminal 1-234aa was identified protection through studying a series of TERT deletion mutant. These findings provide a theoretical base not only for carrying out structure-function study of TERT protecting chromosome end in cancer but also for developing new anti-cancer targets based on telomerase.Conclusion: Our study suggests that telomerase can maintain telomere stability independent of its telomerase activity.TERT silence-induced telomere dysfunction is mediated by the ATR-ATM-CHK2-P53-apopotosis pathwayWe find the 1-234aa domain of TERT N terminal interacts with TPP1 and...