Organization and regulation of the human insulin locus

我们正在对人类胰岛细胞中胰岛素基因位点及其附近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置之间的核内远程物理接触进行测量。为了检测这种相互作用，我们在人的胰岛上进行了4C(一种扩展的染色质构象捕获)实验，以定位核内INS启动子与11号染色体上远程位置之间的联系。我们选择了一个候选基因--Synaptopagmin 8(Syt8)基因，它与胰岛素基因一样，具有邻近的CTCF结合位点，用于进一步研究。我们发现INS-Syt8接触是存在的，它们被添加葡萄糖刺激，而当INS启动子被阻断以沉默INS基因活性时，它们被抑制。接触增多与Syt8表达增加相关。CTCF的耗尽会导致失去联系和Syt8(但不是INS)表达的丧失。此外，我们还发现Syt8的缺失会导致胰岛分泌的胰岛素减少。这些结果揭示了一个远程相互作用的物理网络，这导致了耦合的监管控制。这些类型的调控网络似乎存在于整个基因组中。我们现在正在将我们的4C方法扩展到人类胰岛β细胞系中，精确度大大提高。 我们现在发现了一个有着类似行为的不同网站。Syt8基因座距离INS仅约300kb，而新的基因座距离INS约50Mb，约为11号染色体长度的一半。该新基因座位于与I型糖尿病易感性相关的基因座上。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的分泌，我们正在详细探索这种可能性。

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.