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

项目概要 脂肪组织对于全身能量稳态至关重要。肥胖和脂肪营养不良（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“青少年”，表现出生长迟缓、脂肪营养不良、极端胰岛素抵抗、严重糖尿病 空腹血糖高达600-700 mg/dL，易患脂肪肝。微小的表型是由无效突变引起的 包含 2 (KBTBD2) 的 Kelch 重复序列和 BTB (POZ) 结构域，其没有先前指定的功能。 我们发现KBTBD2作为E3泛素连接酶，通过靶向调节胰岛素信号通路 磷脂酰肌醇 3-激酶 (PI3K) 调节亚基 p85α 的降解。该提案将进一步 探索 KBTBD2 在脂肪营养不良、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调节因子。在指导的K99阶段，将生成小鼠 操纵 Kbtbd2 在不同组织中的表达，研究其组织特异性功能，并剖析其作用 每个组织在微小表型发育中的作用（目标 1）。我发现 KBTBD2 包含一个 YXXM 基序， 它可以被磷酸化以招募 p85α。 KBTBD2 的磷酸化可以解释为什么 p85α 在不同的细胞和组织中选择性地降解。反过来，这表明 KBTDB2 本身可能是 根据组织、发育阶段和/或代谢状态受到翻译后调节。 目标 2 的重点是研究 KBTBD2 的翻译后修饰。在独立R00阶段， 我将进一步探讨观察到的 p85α 单泛素化的生物学后果（目标 3）。的 饮食诱导的肥胖中 Kbtbd2 的转录抑制导致 p85α 的积累，提示我们 寻找去泛素酶，它可能是抑制剂的目标，以降低肥胖者患糖尿病的风险。在 初步研究已鉴定出几种候选 p85α 去泛素酶。识别新基因 与肥胖和糖尿病有关，我们在小鼠中建立了几项正向遗传筛选，发现 有趣的点击。其中，第二种 BTB 蛋白 (RHOBTB2) 可能在我们现有的总体框架内发挥作用 建造的。在目标 4 中，我将研究去泛素酶和其他新发现的肥胖调节因子的机制。 糖尿病。成功完成本提案中概述的目标将提高我们对 KBTBD2 的理解 并可能为糖尿病治疗提供新的靶点。为了完成拟议的研究，我将邀请博士。 Philipp Scherer（UT 西南医学中心）作为我的共同导师，在 K99 指导阶段获得了 更多脂肪细胞和糖尿病研究方面的培训。我的导师 Bruce Beutler 博士建立了一个独特的 推进遗传主动性，能够实时识别致病突变。我将继续努力 从已建立的正向遗传筛选中新发现的肥胖和糖尿病调节因子 独立的R00相。这次培训将使我能够扩展我的专业知识并帮助我过渡到 成功的独立学术研究员。

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.