GLUCOCORTICOID RECEPTOR POST-TRANSLATIONAL MODIFICATIONS IN INSULIN RESISTANCE

摘要： 代谢综合征在全球流行，其特征是胰岛素抵抗、高血压和 中心性肥胖。尽管代谢综合征的病因学仍然是一个活跃的研究领域，但一个领先的 假设它是由关键的胰岛素反应组织中糖皮质激素(GCs)水平升高引起的，如 肝脏和脂肪。GCS是一种由肾上腺产生的应激激素。GCS与GC受体(GR)结合 核激素受体通过多种机制改变靶基因的转录，包括直接 与DNA结合(反式激活)以及蛋白质-蛋白质介导的抑制。GCS也是有效的 治疗许多炎症性疾病，包括慢性阻塞性肺疾病，炎症性 肠道疾病和类风湿性关节炎，但由于不利的代谢影响，其使用受到限制，包括 体重增加、胰岛素抵抗和高血压。最近，几个组织发现了GR POST- 翻译修饰(PTM)，这些PTM事件改变GR转录活性。然而， GR PTM在体内与生理相关的范例中的作用尚不清楚。根据初步数据，我 假设GR PTM，特别是磷酸化，介导了GC对肝脏的复杂影响 活着。这一假设将在三个具体目标中得到验证：1)我们将确定GR磷酸化的作用 对肝细胞代谢和基因表达的影响。这将通过重新添加野生型或磷酸突变体来实现 GR亚型对GR基因敲除肝癌细胞的作用2)我们将确定GR磷酸化在 调节小鼠肝脏的肝代谢、基因表达和GR占位。这将通过以下方式完成 用GR抗体分析代谢、基因表达和染色质免疫沉淀。我们有 利用CRISPR创造了新的GR基因敲打基因突变小鼠。这些S211A鼠标有一把钥匙 磷酸化的丝氨酸残基突变为丙氨酸。我们将确定哪些GC介导了肝脏的变化 脂质代谢、基因表达和GR占有率依赖于磷酸化。3)我们将确定是否 肥胖患者血清中GR磷酸化与肝脏胰岛素抵抗相关 Roux-en-Y胃旁路手术。这将通过对这些患者的肝脏活检进行染色来完成(以及 对照)用磷酸GR特异性抗体。目前正在开发选择性GR调制器，用于 代谢性疾病的治疗以及更安全的抗炎药物。因此，理解 GR PTM在GCs调节作用中的作用将为代谢性疾病的新治疗提供线索 以及常见的炎症性疾病。

Abstract: There is a global epidemic of metabolic syndrome, characterized by insulin resistance, hypertension, and central obesity. Although the etiology of metabolic syndrome is still an active area of investigation, one leading hypothesis is that it is caused by elevated glucocorticoid (GCs) levels in key insulin responsive tissues such as liver and fat. GCs are stress hormones produced by the adrenal gland. GCs bind to the GC receptor (GR) a nuclear hormone receptor that alters transcription of target genes by multiple mechanisms including direct binding to DNA (transactivation) as well as protein-protein mediated repression. GCs are also effective therapies for many inflammatory diseases including chronic obstructive pulmonary disease, inflammatory bowel disease and rheumatoid arthritis, but their use is limited due the adverse metabolic effects including weight gain, insulin resistance and hypertension. Recently, several groups have identified GR post- translational modification (PTM) in cell lines, and these PTM events alter GR transcriptional activity. However, the role of GR PTM in physiologically relevant paradigms in vivo is unknown. Based on preliminary data, I hypothesize that GR PTM, specifically phosphorylation, mediates the complex effects of GC on the liver in vivo. This hypothesis will be tested in three Specific Aims: 1) We will determine the role of GR phosphorylation on hepatocyte metabolism and gene expression. This will be done by adding back wildtype or phosphomutant GR isoforms to GR knockdown hepatoma cells 2) We will determine the role of GR phosphorylation in mediating heaptic metabolism, gene expression and GR occupancy in the livers of mice. This will be done by analyzing metabolism, gene expression and chromatin immunoprecipitation with antibodies to GR. We have created novel GR knockin phosphomutant mice using CRISPRS. These S211A mice have a key phosphorylated serine residue mutated to alanine. We will determine which GC mediated changes in hepatic lipid metabolism, gene expression and GR occupancy are phosphorylation dependent. 3) We will determine if GR phosphorylation is associated with hepatic insulin resistance in samples from obese patients undergoing Roux-en-Y gastic bypass surgery. This will be done by staining liver biopsies from these patients (as well as controls) with phosphoGR specific antibodies. Currently selective GR modulators are being developed for the treatment of metabolic diseases as well as safer anti-inflammatory medications. Therefore, understanding the role of GR PTM in mediating effects of GCs should shed light on novel treatments for metabolic disease as well as common inflammatory conditions.