Organization and regulation of the human insulin locus

人胰岛素基因座的组织和调节

• 批准号: 8741425
• 负责人: Gary Felsenfeld
• 金额: $ 45.93万
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8741425
• 关键词: Affect; Behavior; Beta Cell; Binding Sites; Cell Line; Cell Nucleus; Chromatin; Chromatin Structure; Chromosomes, Human, Pair 11; Coupled; Defect; Distant; Gene Expression; Genes; Genome; Genomics; Glucose; Human; Insulin; Insulin-Dependent Diabetes Mellitus; Islet Cell; Length; Maps; Measurement; Methods; Molecular Conformation; Neighborhoods; Predisposition; Regulation; Site; Susceptibility Gene; histone modification; insulin secretion; islet; promoter; research study; synaptotagmin

We are carrying out studies of the histone modifications over the insulin gene locus and its neighborhood in human islet cells, in an attempt to identify long range regulatory influences that may affect insulin gene expression. We are carrying out measurements of long range physical contacts within the nucleus between the insulin promoter and other genomic sites. To detect such interactions we performed 4C (an extended chromatin conformation capture) experiments in human islets, to map contacts within the nucleus between the Ins promoter and distant sites on chromosome 11. We selected for further study a single candidate, the synaptotagmin 8 (Syt8) gene, which like the insulin locus, has a nearby CTCF binding site. We find that Ins-Syt8 contacts are present, that they are stimulated by addition of glucose and inhibited when the Ins promoter is blocked to silence Ins gene activity. Increased contact is correlated with increased Syt8 expression. Depletion of CTCF results in loss of contact and loss of Syt8 (but not Ins) expression. Furthermore, we show that depletion of Syt8 results in decreased secretion of insulin from the islets. These results reveal a physical network of long range interactions that leads to coupled regulatory control. It seemed likely that these kinds of regulatory networks occur throughout the genome. We are now extending our 4C methods, with greatly increased precision, in a human islet beta cell line. We have now found a different site with similar behavior. Whereas the Syt8 locus is only about 300 kb away from Ins, the new site is about 50 Mb away, about half the length of chromosome 11. This new site is in a locus associated with Type I diabetes susceptibility. Our analysis strongly suggests that defects in the expression or function of the nearby gene may affect insulin secretion, and we are exploring this possibility in detail.

我们正在对人类胰岛细胞中胰岛素基因座及其邻近区域的组蛋白修饰进行研究，试图确定可能影响胰岛素基因表达的长期调控影响。 我们正在对胰岛素启动子和其他基因组位点之间的细胞核内的长距离物理接触进行测量。 为了检测这种相互作用，我们在人类胰岛中进行了 4C（扩展染色质构象捕获）实验，以绘制 Ins 启动子与 11 号染色体上较远位点之间核内的接触图。我们选择了一个候选基因突触结合蛋白 8 (Syt8) 基因进行进一步研究，该候选基因与胰岛素基因座一样，在附近也有一个 CTCF 结合位点。 我们发现存在 Ins-Syt8 接触，它们通过添加葡萄糖而受到刺激，并在 Ins 启动子被阻断以沉默 Ins 基因活性时受到抑制。 接触的增加与 Syt8 表达的增加相关。 CTCF 的耗尽会导致接触丧失和 Syt8（但不是 Ins）表达丧失。 此外，我们发现 Syt8 的消耗会导致胰岛胰岛素分泌减少。 这些结果揭示了导致耦合监管控制的长程相互作用的物理网络。 此类调控网络似乎可能存在于整个基因组中。 我们现在正在人胰岛β细胞系中扩展我们的4C方法，大大提高了精度。 我们现在发现了一个具有类似行为的不同网站。 Syt8 位点距 Ins 仅约 300 kb，而新位点距 Ins 约 50 Mb，约为 11 号染色体长度的一半。这个新位点位于与 I 型糖尿病易感性相关的位点中。 我们的分析强烈表明附近基因的表达或功能缺陷可能会影响胰岛素分泌，我们正在详细探索这种可能性。