Swi/Snf chromatin remodeling activity is required for islet cell development in vivo

Swi/Snf 染色质重塑活性是胰岛细胞体内发育所必需的

• 批准号: 10314133
• 负责人: Rebecca K Davidson
• 金额: $ 3.08万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-01 至 2022-08-23
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10314133
• 关键词: ATAC-seq; ATP phosphohydrolase; Age; Alleles; Alpha Cell; Anatomy; Apoptosis; Area; Automobile Driving; B-Cell Development; Beta Cell; Binding; Biology; Cell Differentiation process; Cell physiology; Cells; Chromatin; Chromatin Remodeling Factor; Core Facility; Data; Development; Diabetes Mellitus; Educational workshop; Embryo; Endocrine; Enhancers; Evaluation; Excision; Faculty; Fasting; Fellowship; Functional disorder; Gene Expression; Gene Expression Profile; Genes; Genetic Transcription; Glucose; Hormone secretion; Hormones; Human; Hyperglycemia; Immunofluorescence Immunologic; Impairment; In Vitro; Indiana; Insulin; Insulin Resistance; Islet Cell; Islets of Langerhans; Mature B-Lymphocyte; Metabolic Diseases; Methods; Modeling; Mus; Mutant Strains Mice; Nature; Non-Insulin-Dependent Diabetes Mellitus; Pancreas; Patients; Peripheral; Phenotype; Physiological; Plasma; Play; Production; Protocols documentation; Regulation; Research; Research Proposals; Role; Series; Stains; Stem Cell Development; Testing; Time; Training; Universities; Weaning; XCL1 gene; blood glucose regulation; career development; cell type; chromatin remodeling; effective therapy; endocrine pancreas development; experience; functional loss; genetic corepressor; glucose tolerance; impaired glucose tolerance; in vivo; islet; islet stem cells; medical schools; mutant; novel; pancreas development; pancreatic islet function; postnatal; pre-doctoral; precursor cell; progenitor; programs; promoter; protein protein interaction; recruit; stem cell differentiation; stem cell proliferation; stem cells; transcription factor; transcriptome sequencing; transdifferentiation

PROJECT SUMMARY: Type 2 diabetes is associated with loss of activity and expression of pancreas-enriched transcription factors (TFs) that control gene expression programs necessary for mature β-cell function. Among these is Pdx1, which plays key roles during early pancreas development, islet endocrine cell differentiation, and β-cell development and function. Pdx1, like most TFs, requires additional regulation from recruited coregulators to modulate specific regulatory programs. Numerous coregulators have been identified as Pdx1-interacting partners including the ATP-dependent Swi/Snf chromatin remodeling complex. In the early developing pancreas, Swi/Snf plays a critical role in progenitor cell expansion, where early pancreatic progenitor cell deletion of one essential Swi/Snf ATPase subunit, Brg1, results in a 50% reduction in final pancreas mass. In the mature β-cell, deletion of both Swi/Snf ATPase subunits, Brg1 and Brm, impairs whole-body glucose tolerance through severe loss of insulin production, which is largely driven by a loss of Pdx1 occupancy on the insulin gene promoter. While Swi/Snf plays an essential role in the early developing pancreas and the mature β-cell, the contribution of Swi/Snf chromatin remodeling activity to islet endocrine progenitor development has not yet been explored. Herein, I will test the hypothesis that the Pdx1-recruited Swi/Snf chromatin remodeling complex dynamically controls the chromatin landscape and expression of genes essential for endocrine progenitor cell development and postnatal islet function. Mutant mice with endocrine-specific deletions of either Brg1, Brm, or both subunits were generated to determine the mechanistic actions of Swi/Snf at this stage of islet development and evaluate the postnatal consequences of losing Swi/Snf during islet endocrine cell development. Preliminary results demonstrate that loss of the Brg1 subunit, but not Brm, from endocrine progenitors leads to severe glucose dyshomeostasis beginning at 4 weeks of age with a reduction in plasma insulin levels, suggesting that Brg1 is essential for proper islet development and function. Remarkably, no mice deficient for both subunits have been recovered at weaning, indicating that total loss of Swi/Snf activity results in postnatal lethality. Aim 1 will explore the anatomical and physiological attributes driving the postnatal phenotype observed in the Swi/Snf mutants through quantitation of islet cell mass and evaluation of islet function through perifusion analysis on isolated islets. Aim 2 will investigate the mechanistic actions of Swi/Snf in controlling chromatin accessibility, TF recruitment, and gene expression programs in endocrine progenitor cells with RNA-sequencing, ATAC-sequencing, and ChIP-qPCR approaches. With this F31 Predoctoral Fellowship, I will be able to commit my time to completing the research described in the Aims outlined in this application, while also focusing on career development and enhancing my scientific skillset through attending seminars and workshops. Indiana University School of Medicine is equipped with experienced faculty and state-of-the-art core facilities to assist in carrying out this research proposal and provide guidance in my training.

项目摘要：2型糖尿病与胰腺富集的活性丧失和表达有关 转录因子（TF）控制成熟β细胞功能所需的基因表达程序。之中 这些是PDX1，它在胰腺早期发育，胰岛内分泌细胞分化和 β细胞的发育和功能。像大多数TF一样，PDX1需要对招聘的核心调节器进行额外的法规 调节特定的监管计划。许多核心节已被确定为PDX1相互作用 包括依赖ATP的SWI/SNF染色质重塑复合物在内的合作伙伴。在早期开发的胰腺中， SWI/SNF在祖细胞膨胀中起关键作用，其中一种早期的胰祖细胞缺失 必需的SWI/SNF ATPase亚基BRG1导致最终胰腺质量降低50％。在成熟的β细胞中， 删除SWI/SNF ATPase亚基，BRG1和BRM，通过严重 胰岛素产生的丧失，这在很大程度上是由于胰岛素基因启动子上PDX1占用率的丧失所致。 SWI/SNF在早期发育的胰腺和成熟的β细胞中起着至关重要的作用，但 尚未探索SWI/SNF染色质重塑活性到胰岛内分泌祖细胞发育。 在此，我将测试以下假设 动态控制内分泌必不可少的基因的染色质景观和表达 祖细胞发育和产后胰岛功能。具有内分泌特定缺失的突变小鼠 生成BRG1，BRM或两个亚基以确定SWI/SNF在此阶段的机械作用 胰岛发育并评估胰岛内分泌细胞中失去SWI/SNF的产后后果 发展。初步结果表明，内分泌的BRG1亚基的丢失，而不是BRM 祖细胞从4周龄开始，血浆降低，导致严重的葡萄糖dyshomeostasis 胰岛素水平，表明BRG1对于适当的胰岛发育和功能至关重要。值得注意的是，没有老鼠 断奶时两个亚基的缺陷都已回收，表明SWI/SNF活动结果的总损失 在产后致死性。 AIM 1将探索驱动产后的解剖学和物理属性 通过定量胰岛细胞质量和胰岛功能评估，在SWI/SNF突变体中观察到的表型 通过对孤立胰岛的渗出分析。 AIM 2将调查SWI/SNF的机械作用 控制内分泌祖细胞中的染色质可及性，TF募集和基因表达程序 有了这个F31冠军奖学金，我 将能够花时间完成本应用程序中概述的目的中描述的研究，而 还专注于职业发展，并通过参加半手体来增强我的科学技能 讲习班。印第安纳大学医学院配备了经验丰富的教师和最先进的核心 协助执行这项研究建议并在我的培训中提供指导的设施。