Telomerase-regulated gene expression in normal and tumor cells

描述（由申请人提供）：端粒酶是通过合成重复端粒DNA来维持染色体末端（端粒）的酶。这防止了染色体复制过程中端粒的侵蚀，从而防止了当端粒变得非常短时发生的遗传不稳定和细胞衰老。然而，一组令人信服的观察结果现在支持这样的假设，即端粒酶具有第二种不依赖端粒的功能，其也促进细胞生长和分裂，抑制细胞死亡，并有助于致癌细胞转化。我们最近的研究首次从机制上了解了端粒酶的这种新功能，表明在人类乳腺上皮细胞中，端粒酶调节一组十二个基因的表达，这些基因可能直接促进细胞增殖并抑制细胞死亡。本课题的具体工作将是了解端粒酶诱导基因表达程序的生物学效应。首先，我们将研究端粒酶调节基因，单独和组合，对乳腺细胞增殖和细胞死亡的影响，并使用三维体外模型上皮腺泡发育，我们将研究其对乳腺形态发生的影响。其次，我们将确定介导端粒酶诱导和抑制基因表达能力的结构域。基于这些研究，我们将构建功能分离突变，这将用于确定端粒的维持和基因表达的调控对细胞存活，衰老和增殖的具体贡献。第三，我们将使用端粒酶突变体和端粒酶调节基因来检验端粒酶对基因表达的调节对于其促进细胞转化的能力是必不可少的这一假设。这些实验将使用二维和三维细胞培养模型，以及致癌转化的转基因小鼠模型。总的来说，这些研究将对我们理解端粒酶独立于端粒维持促进肿瘤发生的机制产生重要影响。这些知识将直接影响我们对端粒酶在癌症中的作用的思考，并且对于开发针对端粒酶和/或其下游信号通路的致癌功能的有效疗法至关重要。

DESCRIPTION (provided by applicant): Telomerase is the enzyme that maintains chromosome ends (telomeres), by synthesizing repetitive telomeric DNA. This prevents telomere erosion during chromosome replication, and thereby prevents the genetic instability and cellular senescence that occurs when telomeres become critically short. However, a compelling set of observations now support the hypothesis that telomerase has a second, telomere- independent function which also promotes cell growth and division, inhibits cell death, and contributes to oncogenic cell transformation. Our recent studies provided the first mechanistic insight into this new function for telomerase, by showing that in human mammary epithelial cells telomerase modulates the expression of a set of twelve genes that may directly promote cell proliferation and inhibit cell death. The specific work of this proposal will be to understand biological effects of the program of gene expression induced by telomerase. First, we will study the effects of telomerase-regulated genes, individually and in combination, on mammary cell proliferation and cell death, and using a three dimensional in vitro model of epithelial acinar development we will study their effects on mammary morphogenesis. Second, we will identify the domains in telomerase that mediate its ability to induce and repress gene expression. Based on these studies we will construct separation-of-function mutations, which will be used to determine the specific contributions that telomere maintenance and regulation of gene expression make to cell survival, senescence and proliferation. Third, we will use telomerase mutants and telomerase-regulated genes to test the hypothesis that modulation of gene expression by telomerase is essential for its ability to promote cell transformation. These experiments will use two- and three-dimensional cell culture models, and transgenic mouse models of oncogenic transformation. Collectively, these studies will have important impact on our understanding of the mechanisms by which telomerase promotes tumorigenesis independently of telomere maintenance. This knowledge would directly impact our thinking about telomerase's role in cancer and will be critical for the development of effective therapies targeting the oncogenic functions of telomerase and/or its downstream signaling pathways.