Investigating imitation SWI chromatin remodeling complexes in mammalian tissue regeneration

研究哺乳动物组织再生中的仿 SWI 染色质重塑复合物

基本信息

批准号：
10030411
负责人：
Hao Zhu
金额：
$ 36.77万
依托单位：
UT SOUTHWESTERN MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-07-01 至 2024-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10030411
关键词：
ATP Hydrolysis Bromodomain CRISPR screen ChIP-seq Chemicals Chromatin Remodeling Factor Clinical Clinical Drug Development Colitis Complex Data Disease Drug Targeting Enzymes Epigenetic Process Excision Genes Genetic Genetic Transcription Genetic Translation Genomics Goals Health Hepatocyte Histones Human In Vitro Inflammatory Bowel Diseases Intestinal Diseases Intestines Knockout Mice Liver Liver Cirrhosis Liver Regeneration Liver diseases Long-Term Effects Malignant Neoplasms Mammals Mediating Modeling Molecular Mus Natural regeneration Newts Nucleosomes Operative Surgical Procedures Output Pathway interactions Pharmacology Phenotype Protein Biosynthesis Proteins Reader Recovery Ribosomal DNA Ribosomal RNA Role SMARCA5 gene SWI/SNF Family Complex System Testing Tissues Zebrafish chromatin remodeling genome-wide in vivo inhibitor/antagonist mouse model organ regeneration regenerative regenerative therapy small molecule small molecule inhibitor therapeutic target tissue regeneration validation studies

项目摘要

PROJECT SUMMARY The potent regenerative capabilities of planaria, newts, and zebrafish are largely lost in mammals, an observation that has a tangible impact on human health. Compromised tissue regeneration likely contributes to disorders such as liver cirrhosis or inflammatory bowel diseases (IBD). Limited regeneration in liver disease settings can also preclude the ability to perform large surgical resections for cancer. The relative lack of regenerative therapies could in part be because discovering new targets is difficult and is dependent on in vivo systems that cannot be replicated in the culture dish. To identify new pathways that can be targeted to enhance regeneration, we established an in vivo CRISPR-Cas9 screening platform to evaluate genes in the mouse liver. Using this high-throughput system, we assessed the impact of 152 genes encoding epigenetic machines such as histone readers, writers, and erasers [2]. This identified two imitation SWI/SNF (ISWI) chromatin remodeling complex subunits encoded by Baz2a and Baz2b, genes that were not previously known to regulate regeneration. The interaction between BAZ2 proteins with the SMARCA5 enzyme defines the nucleolar remodeling complex (NoRC), one of five subtypes of ISWI complexes that uses ATP hydrolysis to remodel nucleosomes, particularly at ribosomal DNA (rDNA) loci [3]. Our validation studies showed that in vivo Cas9 deletion of either Baz2a or Baz2b increased liver regeneration in a hepatocyte repopulation model. Similarly, chemical inhibition of BAZ2A and BAZ2B using the specific bromodomain inhibitor GSK2801 resulted in increased liver regeneration [4]. Moreover, we also found that GSK2801 could promote intestinal recovery in a mouse model of colitis. These data suggest that BAZ2 containing ISWI/NoRC chromatin remodeling complexes are important for organ regeneration and promising therapeutic targets for multiple diseases. Our central hypothesis is that the imitation SWI components BAZ2A and BAZ2B limit efficient tissue regeneration by restricting increases in protein synthesis via suppression of rRNA transcription. We will test different aspects of this hypothesis by validating Baz2a and Baz2b genes as bona fide regeneration regulators (Aim 1), by understanding the global epigenetic activities of BAZ2 containing complexes (Aim 2), and by determining if increased protein synthesis is a key mechanism by which BAZ2 inhibition promotes regeneration (Aim 3). This project will define the regenerative phenotypes and molecular mechanisms associated with ISWI chromatin remodeling. Our studies will facilitate clinical drug development of small molecule inhibitors of BAZ2 proteins for use in enhancing tissue regeneration in the liver and intestine.

项目概要一项观察显示，涡虫、蝾螈和斑马鱼的强大再生能力在哺乳动物中很大程度上丧失了这对人类健康产生了切实的影响。组织再生受损可能导致疾病例如肝硬化或炎症性肠病（IBD）。肝病环境中再生有限可以也排除了对癌症进行大型手术切除的能力。相对缺乏再生能力治疗方法可能部分是因为发现新靶点很困难，并且依赖于体内系统不能在培养皿中复制。为了确定可增强再生的新途径，我们建立了体内 CRISPR-Cas9 筛选平台来评估小鼠肝脏中的基因。使用这个高通量系统，我们评估了 152 个编码表观遗传机器（例如组蛋白）的基因的影响读取器、写入器和擦除器[2]。这鉴定了两个仿 SWI/SNF (ISWI) 染色质重塑复合物由 Baz2a 和 Baz2b 编码的亚基，这些基因以前并不知道调节再生。这 BAZ2 蛋白与 SMARCA5 酶之间的相互作用定义了核仁重塑复合物 (NoRC)，ISWI 复合物的五种亚型之一，利用 ATP 水解来重塑核小体，特别是位于核糖体 DNA (rDNA) 基因座 [3]。我们的验证研究表明，体内 Cas9 删除 Baz2a 或 Baz2b 在肝细胞再生模型中增加肝脏再生。同样，BAZ2A 的化学抑制 BAZ2B 使用特定的溴结构域抑制剂 GSK2801 导致肝再生增加 [4]。此外，我们还发现 GSK2801 可以促进结肠炎小鼠模型的肠道恢复。这些数据表明，含有 ISWI/NoRC 染色质重塑复合物的 BAZ2 对于器官至关重要再生和多种疾病的有希望的治疗靶点。我们的中心假设是模仿 SWI 成分 BAZ2A 和 BAZ2B 通过限制蛋白质的增加来限制有效的组织再生通过抑制 rRNA 转录来合成。我们将通过验证来测试这个假设的不同方面通过了解全局表观遗传，Baz2a 和 Baz2b 基因作为真正的再生调节因子（目标 1）含有复合物的 BAZ2 的活性（目标 2），并确定增加的蛋白质合成是否是关键 BAZ2 抑制促进再生的机制（目标 3）。该项目将定义再生与 ISWI 染色质重塑相关的表型和分子机制。我们的研究将促进用于增强组织再生的 BAZ2 蛋白小分子抑制剂的临床药物开发在肝脏和肠道中。