The Role of KBTBD2 in Lipodystrophy, Insulin Resistance, and Diabetes

KBTBD2 在脂肪营养不良、胰岛素抵抗和糖尿病中的作用

• 批准号: 10468738
• 负责人: Zhao Zhang
• 金额: $ 24.9万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10468738
• 关键词: 1-Phosphatidylinositol 3-Kinase; Adipocytes; Adipose tissue; Affect; Antibodies; BTB/POZ Domain; Binding; Biochemical; Biochemistry; Biological; Biological Assay; Blood Glucose; Body fat; Cells; Clustered Regularly Interspaced Short Palindromic Repeats; Complex; Cre-LoxP; Data; Defect; Development; Diabetes Mellitus; Diabetic mouse; Exhibits; Fasting; Fatty Liver; Fatty acid glycerol esters; Genes; Genetic; Genetic Screening; Genetic Transcription; Glucose; Growth; Homeostasis; Human; In Vitro; Insulin Resistance; Insulin Signaling Pathway; Knock-out; Lipids; Lipodystrophy; Liver; Maintenance; Malignant Neoplasms; Mass Spectrum Analysis; Mediating; Medical center; Mentors; Metabolic; Metabolic Diseases; Modification; Monoubiquitination; Mus; Muscle; Mutant Strains Mice; Mutate; Non-Insulin-Dependent Diabetes Mellitus; Nonsense Mutation; Obesity; Pathway interactions; Phase; Phenotype; Phosphorylation; Phosphotransferases; Physiologic Monitoring; Play; Polyubiquitination; Post-Translational Protein Processing; Post-Translational Regulation; Prevalence; Proteins; Public Health; Regulation; Research; Research Personnel; Role; Site; System; Testing; Tetanus Helper Peptide; Therapeutic; Time; Tissues; Training; Transplantation; Ubiquitination; Undifferentiated; Work; adipocyte differentiation; causal variant; conditional knockout; cullin-3; diabetes pathogenesis; diabetes risk; diet-induced obesity; fasting glucose; global health; improved; in vivo; inhibitor; insight; insulin sensitivity; insulin signaling; metabolic phenotype; mouse genetics; mouse model; new therapeutic target; novel; null mutation; overexpression; protein function; recruit; tool; ubiquitin-protein ligase

PROJECT SUMMARY Adipose tissue is critical for whole body energy homeostasis. Both obesity and lipodystrophy (caused by loss of fat) are associated with diabetes and insulin resistance. Our recent work identified a new mutant mouse, termed teeny, that exhibited growth retardation, lipodystrophy, extreme insulin resistance, severe diabetes with fasting glucose as high as 600-700 mg/dL, and fatty liver. The teeny phenotype is caused by a null mutation in the Kelch repeat and BTB (POZ) Domain containing 2 (KBTBD2), which has no previously assigned functions. We found that KBTBD2 operates as an E3 ubiquitin ligase to regulate the insulin signaling pathway by targeting the degradation of the regulatory subunit of phosphatidylinositol 3-kinase (PI3K), p85α. This proposal will further explore the role of KBTBD2 in lipodystrophy, insulin resistance, and diabetes, and will seek to identify mechanistically related regulators of diabetes and obesity. In the mentored K99 phase, mice will be generated to manipulate Kbtbd2 expression in different tissues to study its tissue-specific functions, and to dissect the role of each tissue in the development of teeny phenotype (Aim 1). I found that KBTBD2 harbors a YXXM motif, which may be phosphorylated in order to recruit p85α. The phosphorylation of KBTBD2 could explain why p85α is selectively degraded in different cells and tissues. This, in turn, would suggest that KBTDB2 itself might be subject to post-translational regulation depending upon tissue, developmental stage, and/or metabolic status. The focus of Aim 2 will be to study the post-translational modification of KBTBD2. In the independent R00 phase, I will further explore the biological consequences of the observed p85α mono-ubiquitination (Aim 3). The transcriptional suppression of Kbtbd2 in diet-induced obesity, leading to the accumulation of p85α, prompts us to search for deubiquitinases, which might be targeted by inhibitors to reduce the risk of diabetes in obesity. In preliminary studies, several candidate p85α deubiquitinases have been identified. To identify new genes that are involved in obesity and diabetes, we have established several forward genetic screens in mice and found interesting hits. Among these, a second BTB protein (RHOBTB2) may act within the overall framework we have built. In Aim 4, I will study the mechanism of deubiquitinases and other newly identified regulators in obesity and diabetes. Successful completion of the aims outlined in this proposal will improve our understanding of KBTBD2 and may provide new targets for diabetes treatment. To accomplish the proposed research, I will include Dr. Philipp Scherer (UT Southwestern Medical Center) as my co-mentor during the mentored K99 phase to gain more training in adipocyte and diabetes research. My mentor, Dr. Bruce Beutler, has established a unique forward genetic initiative with the ability to identify causative mutations in real time. I will continue working on newly identified regulators of obesity and diabetes from the established forward genetic screens in the independent R00 phase. This training will allow me to expand my expertise and help my transition into a successful independent academic researcher.

项目摘要 脂肪组织对整个身体的能量平衡至关重要。肥胖和脂肪代谢障碍（由 脂肪减少）与糖尿病和胰岛素抵抗有关。我们最近的工作发现了一种新的突变小鼠， 被称为teeny，表现出生长迟缓，脂肪代谢障碍，极端胰岛素抵抗，严重的糖尿病， 空腹血糖高达600-700 mg/dL，脂肪肝。微小的表型是由一个无效突变引起的， Kelch重复序列和BTB（POZ）结构域包含2（KBTBD 2），其没有先前指定的功能。 我们发现KBTBD 2作为E3泛素连接酶通过靶向胰岛素信号通路来调节胰岛素信号通路。 磷脂酰肌醇3-激酶（PI 3 K）调节亚基p85α的降解。该提案将进一步 探索KBTBD 2在脂肪代谢障碍、胰岛素抵抗和糖尿病中的作用，并将寻求确定 糖尿病和肥胖症的机制相关的调节剂。在指导的K99阶段，将产生小鼠 操纵Kbtbd 2在不同组织中的表达，以研究其组织特异性功能，并分析其在不同组织中的作用。 每种组织在微小表型发育中的作用（目的1）。我发现KBTBD 2含有YXXM基序， 其可被磷酸化以募集p85α。KBTBD 2的磷酸化可以解释为什么p85α 在不同的细胞和组织中被选择性降解。这反过来又表明，KBTDB 2本身可能是 根据组织、发育阶段和/或代谢状态进行翻译后调节。 目标2的重点是研究KBTBD 2的翻译后修饰。在独立R 00阶段， 我将进一步探讨观察到的p85α单泛素化的生物学后果（目的3）。的 在饮食诱导的肥胖中，Kbtbd 2的转录抑制导致p85α的积累，提示我们 寻找去泛素化酶，这种酶可能被抑制剂靶向，以降低肥胖者患糖尿病的风险。在 初步研究中，已鉴定出几种候选p85α去遍在蛋白酶。去识别新的基因， 与肥胖和糖尿病有关，我们已经在小鼠中建立了几个正向遗传筛选， 有趣的点击。其中，第二种BTB蛋白（RHOBTB 2）可能在我们所知的总体框架内起作用。 建成在目标4中，我将研究去泛素化酶和其他新发现的调节因子在肥胖中的作用机制， 糖尿病成功完成本提案中概述的目标将提高我们对KBTBD 2的理解 并可能为糖尿病治疗提供新的靶点。为了完成这项研究，我将包括博士。 Philipp谢勒（UT西南医学中心）作为我的共同导师在指导K99阶段，以获得 在脂肪细胞和糖尿病研究方面的更多培训。我的导师布鲁斯·博伊特勒博士建立了一种独特的 具有真实的时间识别致病突变的能力的向前遗传主动性。我会继续努力 新发现的肥胖和糖尿病的调节因子， 独立R 00相位。这次培训将使我扩大我的专业知识，并帮助我过渡到一个 成功的独立学术研究者。

项目成果

Generation of epitope tag knock-in mice with CRISPR-Cas9 to study the function of endogenous proteins.

• DOI: 10.1016/j.xpro.2023.102518
• 发表时间: 2023-09-15
• 期刊: STAR PROTOCOLS
• 作者: Zhang, Zhao

Loss of Hilnc prevents diet-induced hepatic steatosis through binding of IGF2BP2.

Hilnc 的缺失通过结合 IGF2BP2 预防饮食诱导的肝脂肪变性

• DOI: 10.1038/s42255-021-00488-3
• 发表时间: 2021-11
• 期刊: NATURE METABOLISM
• 影响因子: 20.8
• 作者: Jiang, Yiao;Peng, Jiayin;Song, Jiawen;He, Juan;Jiang, Man;Wang, Jia;Ma, Liya;Wang, Yuang;Lin, Moubin;Wu, Hailong;Zhang, Zhao;Gao, Dong;Zhao, Yun
• 通讯作者: Zhao, Yun