Elucidating how transcription factors MAFA and MAFB and mitochondrial activity control human β cell identity and function

阐明转录因子 MAFA 和 MAFB 以及线粒体活性如何控制人类细胞的身份和功能

• 批准号: 10660007
• 负责人: Emily M Walker
• 金额: $ 14.25万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-04-01 至 2028-02-29
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10660007
• 关键词: Adult; Affect; B-Lymphocytes; Back; Beta Cell; Biogenesis; Biological Assay; Blood Glucose; Cell Differentiation process; Cell Line; Cell Maturation; Cell physiology; Cells; Complement Factor B; Consumption; Data; Defect; Diabetes Mellitus; Equilibrium; Etiology; Failure; Feedback; Functional disorder; Future; Gene Expression; Gene Expression Profile; Gene Expression Profiling; Gene Expression Regulation; Gene Targeting; Genes; Genetic; Genetic Transcription; Glucose; Health; Homo; Human; Impairment; Insulin; Life Cycle Stages; Metabolic; Metabolic Control; Mitochondria; Mitochondrial DNA; Mus; Non-Insulin-Dependent Diabetes Mellitus; Nuclear; Oxidative Stress; Oxygen Consumption; Pathway interactions; Patients; Phosphotransferases; Physiological; Proteins; Regulation; Reporting; Role; Signal Transduction; Structure; Testing; Transplantation; biological adaptation to stress; blood glucose regulation; cell dedifferentiation; factor A; functional restoration; human embryonic stem cell; humoral immunity deficiency; in vivo; insulin secretion; islet; knock-down; maturity onset diabetes of the young; metabolomics; mitochondrial dysfunction; oxidative damage; pharmacologic; programs; response; single-cell RNA sequencing; stem cells; targeted treatment; tool; transcription factor

Project Summary Type 2 diabetes (T2D) can be attributed to loss of β-cell identity or de-differentiation, marked by acquisition of immature cell markers and loss of insulin expression and secretion. While the etiology of β-cell immaturity in T2D is unclear, impairments in nuclear-encoded mitochondrial gene expression and transcription factor (TF) expression occur. Additionally, defects in mitochondrial structure and function leads to impaired glucose- stimulated insulin secretion (GSIS) and have been reported in β-cells of human T2D patients. Interestingly, the transcriptional changes that occur during β-cell immaturity involve loss of the nuclear-encoded mitochondrial gene expression program. My studies will test the hypothesis that β-cell immaturity in T2D is driven by loss of mitochondrial functional gene regulation by the TFs MAF bZIP transcription factor A (MAFA) or B (MAFB). Furthermore, I predict that MAFA/MAFB are themselves targets of mito-nuclear crosstalk through a retrograde signaling cascade induced by defects in mitochondrial function. I will elucidate the contribution of MAFA and MAFB on metabolic control in human β cells through regulation of mitochondrial function (Aim 1). I will determine how loss of MAFA and/or MAFB affects mitochondrial function and β cell identity by assaying oxygen consumption and gene expression in MAFA and/or MAFB knockdown human pseudoislets and EndoC-βH3 β cell lines. Metabolomics will be performed on EndoC- βH3 β cell lines to determine how MAFA/B influences fuel utilization. My preliminary data shows that genetic loss of mitophagy (i.e., the balance of mitochondrial biogenesis and turnover) reduces β-cell maturity. This includes physiologic, metabolic, and transcriptional signatures consistent with metabolic overload, oxidative damage, and the integrated stress response (ISR). MAFA (and likely MAFB) is known to be more sensitive to oxidative stress associated with T2D β-cell dysfunction than other TFs. While nuclear expression of β-cell mitochondrial genes are well known, mitochondrial feedback to drive β-cell nuclear gene expression (retrograde signaling) has not been analyzed. I will delineate if TF levels and β-cell maturity are altered in response to mitochondrial dysfunction (Aim 2). Further, I observed that pharmacological inhibition of the ISR relieves markers of immaturity in islets of mitophagy-deficient mice. Utilizing pharmacologic tools and analysis of gene expression in human pseudoislet transplants and EndoC-βH3 cells, I will interrogate how such conditions impact human β cells. I expect that MAFA (and possibly MAFB) levels will be reduced because of their ISR sensitivity. Moreover, I will determine if inhibition of the ISR restores MAFA/MAFB expression and reverses β-cell immaturity in the background of mitochondrial damage.

项目摘要 2型糖尿病（T2 D）可归因于β细胞身份的丧失或去分化，其标志为β细胞的获得。 未成熟细胞标记物和胰岛素表达和分泌的丧失。虽然T2 D中β细胞不成熟的病因学 目前尚不清楚，核编码的线粒体基因表达和转录因子（TF） 表达发生。此外，线粒体结构和功能的缺陷导致葡萄糖受损- 刺激的胰岛素分泌（GSIS），并且已经在人T2 D患者的β细胞中报道。有趣的是 在β细胞不成熟期间发生的转录变化涉及核编码的线粒体DNA的丢失， 基因表达程序。我的研究将检验T2 D中β细胞不成熟是由β细胞缺失驱动的假设。 通过TF MAF bZIP转录因子A（MAFA）或B（MAFB）的线粒体功能基因调控。 此外，我预测，MAFA/MAFB本身是通过一个逆行的线粒体核串扰的目标， 线粒体功能缺陷引起的信号级联反应。 我将阐明MAFA和MAFB对人β细胞代谢控制的贡献， 线粒体功能的调节（目的1）。我将确定MAFA和/或MAFB的丢失如何影响 线粒体功能和β细胞特性，通过测定MAFA和/或 MAFB敲低人假胰岛和EndoC-βH3 β细胞系。代谢组学将在EndoC上进行- βH3 β细胞系，以确定MAFA/B如何影响燃料利用。我的初步数据显示基因缺失 线粒体自噬（即，线粒体生物发生和周转的平衡）降低β-细胞成熟。这包括 生理、代谢和转录特征与代谢过载、氧化损伤和 综合应激反应（ISR）。已知MAFA（可能还有MAFB）对氧化应激更敏感， 与T2 D β-细胞功能障碍相关。而β细胞线粒体基因的核表达 众所周知，驱动β细胞核基因表达的线粒体反馈（逆行信号传导）并没有 被分析。我将描述TF水平和β细胞成熟度是否因线粒体功能障碍而改变 (Aim 2）的情况。此外，我观察到ISR的药理学抑制减轻了胰岛中的不成熟标志物， 线粒体自噬缺陷小鼠。利用药理学工具和分析人假胰岛中的基因表达 移植和EndoC-βH3细胞，我将询问这些条件如何影响人类β细胞。我预计 由于ISR敏感性，MAFA（可能还有MAFB）水平将降低。此外，我将确定， 抑制ISR可恢复MAFA/MAFB表达并逆转β细胞不成熟， 线粒体损伤