Insulin Signaling and Metabolic Effects through CLK2 Kinase

CLK2 激酶的胰岛素信号传导和代谢效应

• 批准号: 8638955
• 负责人: Pere Puigserver
• 金额: $ 38.6万
• 依托单位: DANA-FARBER CANCER INST
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-04-01 至 2016-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8638955
• 关键词: Affect; Applications Grants; Binding; Biochemical; Blood Glucose; Buffers; Clinical; Complex; Defect; Diabetes Mellitus; Diabetic mouse; Diet; Disease; Fasting; Gene Expression Profiling; Genes; Genetic; Gluconeogenesis; Glucose; Glycogen; Glycolysis; Goals; Hepatic; Hepatocyte; Homeostasis; Hormones; Hyperglycemia; In Vitro; Insulin; Insulin Resistance; Investigation; Laboratories; Lipids; Liver; Mediating; Mediator of activation protein; Metabolic; Metabolic Control; Metabolic Diseases; Metabolic syndrome; Metabolism; Mitochondria; Molecular; Mus; Non-Insulin-Dependent Diabetes Mellitus; Nutrient; Obesity; Outcome Study; PGC-1 protein; Pathway interactions; Phosphorylation; Phosphorylation Site; Phosphotransferases; Process; Proteins; Proteomics; Regulation; Repression; Signal Transduction; Testing; Tissues; Transcription Coactivator; Translating; adenoviral-mediated; base; blood glucose regulation; db/db mouse; diabetic; fatty acid oxidation; feeding; forkhead protein; genetic analysis; glucose metabolism; glucose output; glucose production; glucose tolerance; glucose transport; hepatic gluconeogenesis; in vivo; insulin signaling; insulin tolerance; lipid metabolism; loss of function; metabolic abnormality assessment; mouse model; mutant; novel; programs; protein complex; protein degradation; public health relevance; research study; response; restoration; small hairpin RNA; transcription factor; ubiquitin ligase

DESCRIPTION (provided by applicant): Nutrient homeostasis is maintained through a complex regulatory network formed by signaling and transcriptional components that control metabolic genes. The liver is one of the key tissues that depending of the physio/pathological conditions buffers whole body nutrient homeostasis. Insulin is one of the most powerful hormones that affect nutrient regulation and clinically, insulin resistance is a hallmark of the metabolic syndrome including type 2 diabetes. As a consequence of insulin resistance one of the metabolic processes that contribute to maintain the diabetic state is hepatic glucose production that is controlled, at least in part, at the transcriptional level. The PI3K/Akt pathway is one of the main effectors of insulin metabolic action. Akt controls expression of metabolic genes through direct phosphorylation and negative regulation of the forkhead transcription factor FoxO1 and coactivators such as PGC-11 and CRCT2, key components of the transcriptional gluconeogenic program. Although Akt can directly mediate this action, there are conditions such as late refeeding or diabetic states where active Akt does not entirely correlate with suppression hepatic glucose production. This indicates that additional key regulatory components, likely kinases, could mediate this repression. Along these lines, we have recently identified Cdc2-like kinase 2 (Clk2) as a novel component downstream of insulin/Akt and functions as part of the hepatic feeding response. Notably, Clk2 controls expression of gluconeogenic genes, hepatic glucose output and blood glucose levels. Moreover, obese/diabetic db/db mice have lower amounts of Clk2 protein and restoration of the levels corrects hyperglycemia. This result suggests that Clk2 might be dysregulated in conditions of obesity/diabetes and contributes to the clinical manifestations. Based on these findings, the major goal of this proposal is to identify the molecular mechanisms by which insulin controls Clk2 kinase activity and to test Clk2 metabolic functionality in-vitro and in in-vivo mouse models. We have three Specific Aims: Aim 1 is to perform molecular and functional analysis of how insulin controls Clk2 kinase activity focusing on regulation of Clk2 protein degradation through ubiquitin ligases. Aim 2 is devoted to carry out molecular and functional analysis of Clk2- mediated suppression of hepatic gluconeogenesis. We focus on the Clk2-induced phosphorylation and repression of PGC-11. Aim 3 will determine the effects of Clk2 on hepatic glucose and lipid metabolism in mice through loss-of-function of hepatic Clk2 in fasting/feeding cycles and genetic and diet-induced obesity. The outcomes of these studies will provide the identification of the molecular mechanisms by which insulin controls glucose and lipid metabolic effects through Clk2 kinase. Since insulin resistance and as a consequence increased and uncontrolled hepatic glucose output is a major defect that occurs in type 2 diabetes, our investigation on the regulation of Clk2 and PGC-11 might translate into potential therapies to treat this disease.

描述（由申请人提供）：营养物质稳态是通过控制代谢基因的信号传导和转录成分形成的复杂调节网络来维持的。肝脏是根据生理/病理条件缓冲全身营养稳态的关键组织之一。胰岛素是影响营养调节的最强大的激素之一，临床上，胰岛素抵抗是包括 2 型糖尿病在内的代谢综合征的标志。由于胰岛素抵抗，有助于维持糖尿病状态的代谢过程之一是肝葡萄糖的产生，其至少部分地在转录水平上受到控制。 PI3K/Akt 通路是胰岛素代谢作用的主要效应器之一。 Akt 通过叉头转录因子 FoxO1 和 PGC-11 和 CRCT2 等共激活因子（转录糖异生程序的关键组成部分）的直接磷酸化和负调节来控制代谢基因的表达。尽管 Akt 可以直接介导这种作用，但在某些情况下，例如延迟再进食或糖尿病状态，活性 Akt 并不与抑制肝葡萄糖产生完全相关。这表明其他关键调节成分（可能是激酶）可以介导这种抑制。沿着这些思路，我们最近发现 Cdc2 样激酶 2 (Clk2) 是胰岛素/Akt 下游的一种新成分，并作为肝脏进食反应的一部分发挥作用。值得注意的是，Clk2 控制糖异生基因、肝葡萄糖输出和血糖水平的表达。此外，肥胖/糖尿病 db/db 小鼠的 Clk2 蛋白含量较低，恢复该水平可以纠正高血糖。这一结果表明，Clk2 在肥胖/糖尿病情况下可能失调，并导致临床表现。基于这些发现，该提案的主要目标是确定胰岛素控制 Clk2 激酶活性的分子机制，并在体外和体内小鼠模型中测试 Clk2 代谢功能。我们有三个具体目标：目标 1 是对胰岛素如何控制 Clk2 激酶活性进行分子和功能分析，重点是通过泛素连接酶调节 Clk2 蛋白降解。目标 2 致力于对 Clk2 介导的肝糖异生抑制进行分子和功能分析。我们重点关注 Clk2 诱导的 PGC-11 磷酸化和抑制。目标 3 将通过禁食/进食周期中肝脏 Clk2 功能的丧失以及遗传和饮食诱导的肥胖来确定 Clk2 对小鼠肝脏葡萄糖和脂质代谢的影响。这些研究的结果将确定胰岛素通过 Clk2 激酶控制葡萄糖和脂质代谢作用的分子机制。由于胰岛素抵抗以及由此导致的肝葡萄糖输出增加和不受控制是 2 型糖尿病中发生的一个主要缺陷，因此我们对 Clk2 和 PGC-11 调节的研究可能会转化为治疗这种疾病的潜在疗法。