The function of Topoisomerase I SUMOylation in transcription and chemoresistance

项目摘要 拓扑异构酶I（TOP1）是FDA批准的晚期癌症的治疗靶点，最近扩大了其在癌症治疗中的应用。 治疗潜力典型的TOP1毒物，如拓扑替康，通过阻止细胞分裂来杀死快速分裂的细胞。 在拓扑异构酶反应期间从DNA释放TOP1以松弛超螺旋DNA，导致DNA 休息.然而，最近发现TOP1毒素也会改变与自闭症相关的基因表达。 谱系障碍（ASD）和急性炎症反应等，通过一种机制， 独立于DNA损伤。事实上，除了放松超螺旋DNA，TOP 1还调节mRNA 通过与RNA聚合酶II（RNAPII）和剪接因子的相互作用，进行剪接和基因表达。 因此，调节TOP 1转录功能而不引起DNA损伤的新药可以 大大增加TOP1的治疗范围，为疑难疾病提供治疗选择。然而，在这方面， TOP1与mRNA调控的联系机制尚未完全建立。我们最近发现 TOP1的赖氨酸（K）残基391和436的SUMO化调节其转录功能，提供了一种新的转录途径。 能加速机械学进步的重要见解由于这一新的见解，我们处于有利地位， 揭示TOP1和mRNA调控的详细分子机制。该提案将首先测试一个 TOP1 K391/K436 SUMO化调节RNAPII运动和RNAPII偶联的新假说 和剪接体以促进基因转录（Aim 1）。我们将评估转录景观， 剪接模式，包括依赖于TOP 1 K391/K436的剪接变体的存在 SUMO化。我们将确定其他的细胞过程，可能会改变抑制的TOP 1 转录功能，以揭示其用于新疾病治疗的全部潜力（目的2）。我们还将开发一个 新的分子策略来破坏TOP 1对mRNA调控的影响。这一新战略将有助于 用于多种疾病的药物开发，包括炎症诱导的脓毒症和ASD。最后在 除了促进转录外，K391/K436 SUMO化还抑制TOP1拓扑异构酶活性。 因为对TOP1毒物的敏感性与TOP1拓扑异构酶活性水平正相关， 细胞，我们将测试一种令人兴奋的可能性，即通过这种SUMO化暂时阻断TOP 1 SUMO化， 抑制剂在癌症化疗期间可使细胞对TOP1毒物超敏（Aim 3）。

Project Summary Topoisomerase I (TOP1) is a FDA-approved therapeutic target for late-stage cancers with a recently expanded therapeutic potential. Canonical TOP1 poisons, such as topotecan, kill rapidly dividing cells by preventing the release of TOP1 from DNA during its topoisomerase reaction to relax supercoiled DNA, leading to DNA breaks. However, it was recently discovered that TOP1 poisons also alter gene expression linked to autism spectrum disorders (ASD) and acute inflammatory response, among others, via a mechanism that is independent of DNA damage. Indeed, in addition to relaxing supercoiled DNA, TOP1 also regulates mRNA splicing and gene expression through its interaction with RNA polymerase II (RNAPII) and splicing factors. Therefore, new drugs that modulate the transcriptional function of TOP1 without causing DNA damage could greatly increase the therapeutic scope of TOP1, and provide treatment options for difficult diseases. However, the mechanisms linking TOP1 to mRNA regulation are not well established. Our recent discovery that SUMOylation of the lysine (K) residues 391 and 436 of TOP1 regulates its transcriptional function provides a crucial insight that will accelerate mechanistic advances. Due to this new insight, we are well-positioned to uncover the detailed molecular mechanism linking TOP1 and mRNA regulation. This proposal will first test a novel hypothesis that TOP1 K391/K436 SUMOylation regulates RNAPII movement and the coupling of RNAPII and spliceosome to facilitate gene transcription (Aim 1). We will evaluate the transcriptional landscape and splicing patterns, including the presence of splice variants that are dependent on TOP1 K391/K436 SUMOylation. We will identify additional cellular processes that may be altered by the inhibition of the TOP1 transcriptional function to uncover its full potential for new disease treatments (Aim 2). We will also develop a novel molecular strategy to disrupt the effects of TOP1 on mRNA regulation. This novel strategy will facilitate drug development for a wide range of disorders, including inflammation-induced sepsis and ASD. Finally, in addition to facilitating transcription, K391/K436 SUMOylation also suppresses TOP1 topoisomerase activity. Because the sensitivity to TOP1 poisons positively correlates with the level of TOP1 topoisomerase activity in cells, we will test an exciting possibility that transiently blocking TOP1 SUMOylation by this SUMOylation inhibitor could hypersensitize cells to TOP1 poisons during cancer chemotherapy (Aim 3).