The mRNA splicing factor RBM5: A new therapeutic target for TBI

mRNA剪接因子RBM5：TBI的新治疗靶点

基本信息

批准号：
8749903
负责人：
TRAVIS C JACKSON
金额：
$ 19.22万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-07-01 至 2016-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8749903
关键词：
Acute Brain Injuries Anthraquinones Apoptotic Applications Grants Behavioral Brain Brain Diseases Brain Injuries CASP8 and FADD-like apoptosis regulating protein CASP8 gene Caspase Cause of Death Cell Death Cells Cellular Stress Cessation of life Clinical Trials Cognitive Data Drosophila pros protein Drug Targeting Enhancers Enzymes Excision Exclusion Exons Genes Hippocampus (Brain)Human Hydrogen Peroxide Impairment In Vitro Injury Intervention Introns Knowledge Learning Length Lesion Mechanics Mediating Memory Messenger RNA Modeling Mus Neurons Nuclear Nuclear Proteins Pharmaceutical Preparations Play Population Process Protein Isoforms Protein Splicing Proteins Publications RNA Binding RNA Recognition Motif RNA Splicing Rattus Recovery Reporting Role Site Spliceosome Assembly Pathway Spliceosomes Staurosporine Stretching Structure Sulfonic Acids TNF gene TNFSF10 gene TNFSF6 gene Testing Testis Therapeutic Toxic effect Transcript Traumatic Brain Injury Up-Regulation Variant Zinc Fingers cancer cell caspase-2 central nervous system injury controlled cortical impact disability in vitro testing in vivo inhibitor/antagonist injured intravenous administration mRNA Expression mRNA Precursor new therapeutic target novel overexpression pro-caspase-2 protein B protein expression public health relevance response small molecule social

项目摘要

PROJECT SUMMARY Reports estimate that over 90% of all human genes are subject to mRNA splicing. RNA splicing occurs in highly specialized structures termed nuclear spliceosomes. Spliceosomes consist of a few key proteins that catalyze the excision of target introns/exons, plus 100's of co-factors that regulate the activity/efficiency of these enzymes. Spliceosome co-factors direct many facets of splicing dynamics including distribution of different protein variants throughout the body. Brain injury and disease disturb splicing. Pathological changes in spliceosome mechanics alter splice variant expression of different survival and death proteins. An upregulation of maladaptive protein variants may exacerbate neuronal death and impair CNS recovery. RNA Binding Motif 5 (RBM5) is a splicing co-factor. It is highly expressed in the CNS and testis but its function in the healthy or injured brain is unknown. RBM5 briefly localizes to the spliceosome, very early during spliceosome assembly around pre-mRNA targets. A zinc finger domain (RanBP2-Type), and several other RNA binding domains, coordinate its highly selective interaction with spliceable exons in pre-mRNA targets. In cancer cells, RBM5 regulates splice variant selection of caspase-2 (pro-death) and c-FLIP (pro-survival) genes. It promotes the exclusion of exon 9 from caspase-2, and exon 7 from c-FLIP mRNA. This induces caspase2L and c-FLIPL protein expression, respectively. Caspase2L is a pro-death splice variant. The c-FLIPL splice variant also has pro-apoptotic functions. In contrast, RBM5 inhibition causes exon 9/7 retention, respectively, and forces caspase-2s/c-FLIPs expression. Caspase-2s and c-FLIPs are both potent pro-survival variants. Thus RBM5 promotes cell death by upregulating the ratio of pro-death to pro-survival splice variants. RBM5 has not been studied in the brain. High expression in the CNS suggests that it may play a key role in splicing-mediated cell death mechanisms. To the best of our knowledge, no drug has ever been developed to specifically target pro-death splicing mechanisms in the brain - thus the therapeutic value of splicing directed therapies is completely unknown. Our preliminary data show that RBM5 is highly enriched in the hippocampal CA3 formation - a neuron population that is especially vulnerable to traumatic brain injury (TBI). Furthermore, we show that anthraquinone-2-sulfonic acid (AQ2S), the world's first small-molecule RBM5 inhibitor (that blocks the RanBP2-Type domain), decreases neuronal death after TBI in vitro and in vivo. This grant proposal seeks to test our hypothesis that RBM5 is a potent pro-death splicing factor in rat/human neurons (i.e. show that it upregulates caspase-2L and c-FLIPL splicing in these cells), and further confirm our preliminary data showing that AQ2S is the first viable drug to block pro-death splicing mechanisms in the brain.

项目概要报告估计，超过 90% 的人类基因都会受到 mRNA 剪接的影响。 RNA剪接发生在称为核剪接体的高度专业化结构中。剪接体由一些关键蛋白质组成催化目标内含子/外显子的切除，加上数百个调节活性/效率的辅助因子这些酶。剪接体辅助因子指导剪接动力学的许多方面，包括剪接体的分布全身不同的蛋白质变体。脑损伤和疾病会干扰剪接。病变在剪接体机制中，改变不同存活和死亡蛋白的剪接变体表达。一个适应不良蛋白变异的上调可能会加剧神经元死亡并损害中枢神经系统的恢复。 RNA 结合基序 5 (RBM5) 是一种剪接辅助因子。它在中枢神经系统和睾丸中高度表达，但其健康或受伤大脑的功能尚不清楚。 RBM5 短暂定位于剪接体，非常早围绕前 mRNA 靶标的剪接体组装。一个锌指结构域（RanBP2-Type），以及其他几个 RNA 结合域协调其与前 mRNA 靶标中的可剪接外显子的高度选择性相互作用。在癌细胞中，RBM5 调节 caspase-2（促死亡）和 c-FLIP（促生存）的剪接变体选择基因。它促进从 caspase-2 中排除外显子 9，从 c-FLIP mRNA 中排除外显子 7。这导致 caspase2L 和 c-FLIPL 蛋白分别表达。 Caspase2L 是一种促死亡剪接变体。 c-FLIPL 剪接变体还具有促凋亡功能。相反，RBM5 抑制会导致外显子 9/7 保留，分别，并强制 caspase-2s/c-FLIPs 表达。 Caspase-2s 和 c-FLIPs 均具有有效的促生存作用变体。因此，RBM5通过上调促死亡与促存活剪接变体的比率来促进细胞死亡。 RBM5 尚未在大脑中进行研究。 CNS 中的高表达表明它可能发挥关键作用剪接介导的细胞死亡机制中的作用。据我们所知，迄今为止还没有药物专门针对大脑中的促死亡剪接机制而开发 - 因此具有治疗价值剪接定向疗法是完全未知的。我们的初步数据表明 RBM5 高度富集于海马 CA3 结构 - 特别容易受到创伤性脑损伤的神经元群（创伤性脑损伤）。此外，我们还发现了世界上第一个小分子蒽醌-2-磺酸（AQ2S） RBM5 抑制剂（阻断 RanBP2 型结构域）可在体外和体内减少 TBI 后的神经元死亡。该拨款提案旨在检验我们的假设，即 RBM5 是大鼠/人类中有效的促死亡剪接因子神经元（即显示它上调这些细胞中的 caspase-2L 和 c-FLIPL 剪接），并进一步证实我们的初步数据表明，AQ2S 是第一种阻断大脑促死亡剪接机制的可行药物。