| The end replication problem of linear chromosomes leads to the erosion of telomeric DNA with each cell division. A mechanism to counteract telomeric attrition involves the activity of the specialized ribonucleoprotein telomerase. Expression of the catalytic subunit of telomerase, hTERT, is tightly regulated and this regulation is achieved through both genetic and epigenetic mechanisms. Activators such as c-MYC and SP1 and repressors such as MAD1, Menin, and p53 mediate hTERT transcription. Epigenetic control mediated by DNA methylation and histone modifications such as methylation and acetylation can reversibly and potently regulate telomerase activity. Not only is there synergism between the epigenetic regulatory mechanisms of DNA methylation and histone acetylation, there is also significant interaction between the genetic and epigenetic methods of hTERT gene regulation. Understanding mechanisms that govern hTERT expression can lead to insights into cellular processes such as embryonic development, differentiation, cancer progression, and aging because of the integral role of telomerase activity in these occurrences. The studies outlined in this dissertation attempt to elucidate some of the mechanisms modulating hTERT transcription and telomerase activity in the initiation of cellular differentiation and in a human breast cancer model. To this end, we have developed a novel model system to investigate differentially-expressed genes during the course of in vitro human embryonic stem cell differentiation. We have also presented data showing a stronger correlation between hTERT expression and expression of the epigenetic regulators DNMT3a and DNMT3b than between other epigenetic regulatory mechanisms. In the breast cancer model, we have investigated the effects of prolonged all-trans retinoic acid exposure on these cells and have demonstrated changes in cellular morphology, growth rate, and anchorage-independence. We have also observed that this retinoid slowly decreases telomerase activity, possibly due to the epigenetic changes induced at the hTERT promoter. The result of these investigations is a deeper comprehension of the complexities involved in telomerase regulation in normal regulated and unregulated cancer systems. This understanding gives insight into the early events of aging and telomere shortening and could lead to improved anti-cancer therapeutics that utilize modes of hTERT down-regulation to stop the growth of telomerase-addicted neoplastic cells. |
