Mechanisms of subtelomere recombination in telomerase deficient tumors

端粒酶缺陷肿瘤中亚端粒重组的机制

• 批准号: 8320132
• 负责人: TAMMY A MORRISH
• 金额: $ 13.48万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-15 至 2012-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8320132
• 关键词: Accounting; B-Cell Lymphomas; Biological Assay; Biology; Brain Neoplasms; Cell Culture Techniques; Cells; Characteristics; DNA; DNA Repair; DNA biosynthesis; Drosophila genus; Enzymes; Event; Genes; Genetic Recombination; Glioblastoma; Goals; Growth; Human; Human Genome; Length; Loss of Heterozygosity; Mammalian Cell; Mesenchymal; Mobile Genetic Elements; Monitor; Mus; Neurons; Phase; Primary Neoplasm; RNA; Retrotransposition; Retrotransposon; Reverse Transcription; Role; Telomerase; Telomerase RNA Component; Telomerase inhibition; Telomere Maintenance; Testing; Yeasts; gene function; mouse genome; neoplastic cell; novel; repaired; sarcoma; small hairpin RNA; telomere; tumor; tumor growth; tumorigenesis

DESCRIPTION (provided by applicant): Telomere maintenance in the absence of telomerase can occur by various mechanisms, collectively termed ALT, for alternative lengthening mechanisms. Nearly 10% of human tumors, often sarcomas or glioblastomas, lack the enzyme telomerase for telomere maintenance. In order to study these non- telomerase telomere maintenance mechanisms we are using E5myc+mTR-/- mice, genetically deleted for the RNA component of telomerase (mTR), and crossed to E5myc+ mice, which develop B-cell lymphoma. To test the role of recombination in these non-telomerase telomere maintenance mechanisms we are currently using an shRNA approach to inhibit various recombination genes, and then examining the change in growth rate of tumors. To understand more about the mechanisms we then assay the tumors for subtelomere recombination. In this proposal we intend to examine the contribution of genes involved in replication fork stalling, and the role of DNA repair by a novel mechanism known as break-induced replication. To monitor break-induced replication in mammalian cells, we will develop assays to detect the hallmarks of break-induced replication. These characteristics include loss of heterozygosity, non-reciprocal translocations, and segmental duplications. Finally during the independent phase we intend to examine the role of LINE-1 retrotransposition during break-induced replication and telomere maintenance. Non- LTR retrotransposons, referred to as LINE-1 or L1, account for a significant fraction of the mouse and human genomes. These mobile genetic elements move by an RNA intermediate using a mechanism called target-primed reverse transcription. In addition we found that retrotransposition can also occur at endogenous DNA breaks and dysfunctional telomeres. Furthermore, non-LTR retrotransposons in Drosophila entirely contribute to the mechanism of telomere maintenance. Thus we intend to examine whether non-LTR retrotransposons also contribute to telomere maintenance in both human and mouse cells and thus account for the occurrence of tumors lacking telomerase.

描述（由申请人提供）：在没有端粒酶的情况下，端粒的维持可以通过各种机制发生，统称为 ALT，用于替代的延长机制。近 10% 的人类肿瘤（通常是肉瘤或胶质母细胞瘤）缺乏维持端粒的端粒酶。为了研究这些非端粒酶端粒维持机制，我们使用 E5myc+mTR-/- 小鼠，该小鼠从基因上删除了端粒酶 (mTR) 的 RNA 成分，并与 E5myc+ 小鼠杂交，后者发展为 B 细胞淋巴瘤。为了测试重组在这些非端粒酶端粒维持机制中的作用，我们目前正在使用shRNA方法来抑制各种重组基因，然后检查肿瘤生长速率的变化。为了更多地了解该机制，我们随后检测肿瘤的亚端粒重组。在本提案中，我们打算研究参与复制叉停滞的基因的贡献，以及通过一种称为断裂诱导复制的新机制进行 DNA 修复的作用。为了监测哺乳动物细胞中断裂诱导的复制，我们将开发检测断裂诱导复制标志的方法。这些特征包括杂合性丧失、非相互易位和片段重复。最后，在独立阶段，我们打算研究 LINE-1 逆转录转座在断裂诱导的复制和端粒维持过程中的作用。非 LTR 逆转录转座子，称为 LINE-1 或 L1，占小鼠和人类基因组的很大一部分。这些可移动的遗传元件使用一种称为靶标引发的逆转录的机制通过 RNA 中间体移动。此外，我们发现逆转录转座也可能发生在内源性 DNA 断裂和功能失调的端粒上。此外，果蝇中的非 LTR 逆转录转座子完全有助于端粒维持机制。因此，我们打算检查非LTR反转录转座子是否也有助于人类和小鼠细胞中端粒的维持，从而解释缺乏端粒酶的肿瘤的发生。