2023-01-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/646828端粒酶是由RNA與蛋白質複合體所組成，主要的功能是可以將染色體的端粒延長。它主要由 反轉錄酶TERT進行反應 ，利用端粒酶RNA當模板將端粒DNA加到染色體末端。端粒酶的表現與活性可促進細胞增生、癌症形成與幹細胞複製等等。本篇的研究著重於端粒酶RNA與端粒酶活性無關的功能上。端粒酶RNA基因的突變會導致許多人類的疾病，如 先天性角化不良症、再生障礙性貧血等。這些病人常伴隨著骨髓幹細胞的喪失、免疫的缺失與較短的染色體端粒等症狀。前人的研究著重於端粒酶RNA在延長端粒的功能，有趣的是，在最新的研究發現端粒酶RNA可能有扮演更多與端粒延長不相關的功能，這些機制都尚未了解。在本篇的研究中，我們純化RNA與蛋白質複合體的技術，發現端粒酶RNA在細胞中會與調控基因表現的蛋白質有交互作用，也能與參與細胞凋亡機制的蛋白質結合，並且端粒酶RNA能座落到非端粒的染色質上。我們推測TERC功能像是核內非編碼RNA一樣，能與染色質上的蛋白質作用，來影響基因表現與表觀遺傳的調控，進而促進細胞增生與阻止細胞凋亡等等現象。這個研究有三個目標：第一，研究端粒酶RNA如何調控表觀遺傳因子與影響染色質的變化。第二，端粒酶RNA與 CCAR1 蛋白質結合來調控細胞凋亡。第三，找尋端粒酶RNA與這些蛋白質結合的區域與序列。這個計畫具有創新性，因為現今科學界對於端粒酶RNA在表觀遺傳的影響與基因調控都不清楚，這個計畫的執行將可針對上述議題加以研究，找出其中機制，期許對於人類疾病與癌症的治療上有貢獻。 Telomerase is a ribonucleoprotein complex that adds telomeric DNA to the ends of linear chromosomes. It is composed of two core components: the RNA component, TERC, which provides the template for DNA synthesis, and the reverse transcriptase protein component, TERT. The expression of telomerase promotes cell proliferation, tumorigenesis, and stem cell survival and renewal. TERC mutations were identified in patients with dyskeratosis congenita and aplastic anemia displaying hematopoietic, immune defects, and shorter telomeres. Although the major function of TERC is believed to regulate the telomere lengthening, recent studies have shown that TERC can act as a non-coding RNA that protects from apoptosis and regulates other cellular processes independent of its function in telomerase enzymatic activity. However, the molecular mechanism is poorly understood. We have identified TERC interacting proteins in mouse embryonic stem cells using identification of direct interacting protein (iDRiP) assay, and found that TERC binds to proteins involved in rRNA synthesis, transcription regulation, cell cycle and telomere maintenance. Moreover, TERC-ChIRP (Chromatin Isolation by RNA-purification)-seq revealed thousands of TERC genomic binding sites, which are enriched in promoters, introns and exons. We found that TERC target genes are involved in cancer pathways, adherens junction, hedgehog signaling pathway, TGF-beta signaling pathway, ribosomal biogenesis, and inflammatory regulation, suggesting that TERC plays a role beyond telomere maintenance. The objective of this proposal is to dissect the role of TERC RNA in apoptosis and epigenetic regulation. The central hypothesis is that TERC associates with transcriptional regulators to regulate gene expression, as well as interacts with CCAR1 to inhibit apoptosis. In this project, we will test the following hypotheses: TERC binds to chromatin and functions in epigenetic regulation (Aim 1). TERC regulates apoptosis through its interaction with CCAR1 (Aim 2). Mapping the binding motifs of TERC that are associated with its interacting proteins (Aim 3). The proposed research is innovative because the noncanonical function of TERC remains elusive, and our proposed approaches will discover the molecular mechanism of TERC in epigenetic regulation that controls cell proliferation and apoptosis.端粒酶RNA;表觀遺傳;染色質;細胞凋亡;端粒;端粒酶;非編碼RNA;telomerase RNA component; epigenetics; chromatin; apoptosis; telomere; telomerase; long non-coding RNA