Mechanisms of genomic instability, tumor initiation and progression following the disruption of the RTF2-RNase H2 axis

项目概要/摘要 核糖核酸（RNA）在基因组中的异常掺入或保留是基因组遗传缺陷的常见原因。 不稳定性，使其易于水解和下游诱变。RNase H2是一种 通过移除这些基因组来防止基因组不稳定的主要机制- 嵌入的RNA。 我们的实验室最近发现了一种新的调节RNase H2的机制， 终止因子2（RTF 2）参与定位和调节复制叉处的RNA酶H2的水平。 需要进一步阐明这种相互作用，以了解调控背后的基本生物学， RNase H2如何防止基因组不稳定性。有趣的是，RNase H2的拷贝数丢失通常是 在慢性淋巴细胞白血病（CLL）中发现，超过30%的病例。在我的初步工作中，我开发了 各种细胞模型，其中RNA酶H2和RTF 2可以耗尽，我已经表达和纯化 重组RNA酶H2和RTF 2，允许RNA酶H2活性被调节的两种体内研究 通过RTF 2和体外研究它们的相互作用。此外，我还开发了一种新的测定方法， 人细胞基因组中核糖核苷酸掺入的定量分析。本试验将用于 研究RTF 2对RNase H2的调节，并将评估其预测CLL对以下反应的适用性： PARP抑制剂。肿瘤进展背后的RNase H2丢失的直接机制尚未研究。 在这个建议中，基于我上述的初步工作，我将测试RTF 2直接与 并在复制叉处调节RNase H2的活性，并研究RNA酶H2丢失背后的机制。 RNase H2损害基因组稳定性并导致肿瘤进展。 我是威尔康奈尔/洛克菲勒/斯隆凯特琳三机构计划的MD/博士生，我在那里 在洛克菲勒大学的Agata Smogorzewska博士的实验室里。我的长期目标是成为 医生科学家，担任血液肿瘤学家并经营独立的基础科学实验室 作为一个学术性的大学医院。本提案中概述的计划，沿着以下人员的支持和指导 博士Agata Smogorzewska，我的论文研究委员会和三机构MD-PhD计划将帮助我 实现这些职业抱负。

Project Summary/Abstract The aberrant incorporation or retention of ribonucleic acids (RNAs) in the genome is a common cause of genomic instability, rendering it susceptible to hydrolysis and downstream mutagenesis. The enzyme RNase H2 is one of the primary mechanisms protecting against this destabilization of the genome by removing these genome- embedded RNAs. Our lab recently uncovered a novel mechanism of regulation of RNase H2, by uncovering that replication termination factor 2 (RTF2) is involved in localizing and regulating the levels of RNase H2 at the replication fork. Further elucidation of this interaction is required to understand the basic biology behind the regulation and function of how RNase H2 prevents genome instability. Interestingly, copy number loss of RNase H2 is commonly found in Chronic Lymphocytic Leukemia (CLL), in over 30% of cases. In my preliminary work, I have developed various cellular models in which RNase H2 and RTF2 can be depleted, and I have expressed and purified recombinant RNase H2 and RTF2, allowing for both in vivo studies of which RNase H2 activities are regulated by RTF2 and in vitro studies of their interaction. Furthermore, I have developed a novel assay allowing a quantitative analysis of ribonucleotide incorporation in the genomes of human cells. This assay will be used to study the regulation of RNase H2 by RTF2, and will be assessed in its applicability to predict CLL responses to PARP-inhibitors. The direct mechanism behind tumor progression in the loss of RNase H2 has not been studied. In this proposal, building on my above preliminary work, I will test the hypothesis that RTF2 interacts directly with and regulates the activities of RNase H2 at the replication fork and examine the mechanism behind how loss of RNase H2 compromises genomic stability and leads to tumor progression. I am an MD/PhD student at the Weill Cornell/Rockefeller/Sloan Kettering Tri-Institutional Program, where I am in the laboratory of Dr. Agata Smogorzewska at The Rockefeller University. My long-term goal is to become a physician scientist, practicing as a hematologist-oncologist as well as running an independent basic science lab as an academic university hospital. The plan outlined in this proposal, along with the support and mentorship of Dr. Agata Smogorzewska, my thesis research committee, and the Tri-Institutional MD-PhD program will help me achieve these career aspirations.