HNF1A in human endocrine cell development and function

糖尿病是一种复杂的疾病，主要有三种亚型：I型、II型和单基因型。I型是一种自身免疫性疾病，其特征是分泌胰岛素的β细胞缺失。II型是一种代谢紊乱，以胰岛素反应性差和最终的β细胞耗竭和损失为特征。单基因形式是由一些基因的突变引起的，这些基因通常在细胞发育和/或功能中很重要。本提案将重点研究HNF1A，这是一种转录因子，当突变时导致单基因糖尿病最常见的原因。此外，在GWAS研究中，HNF1A与I型和II型糖尿病都有关联。HNF1A缺乏的小鼠模型是可用的，但不幸的是不能适当地模拟人类疾病。因此，使用人类模型系统很重要。我们将利用人多能干细胞模型研究HNF1A在内分泌细胞发育和功能中的作用，利用基于CRISPR-Cas9的基因组工程产生等基因突变系。稳健的体外分化方案存在，并将用于在人类干细胞模型中产生β细胞。在初步研究中，我们发现了人类特异性的HNF1A靶点，这些靶点已在来自HNF1A突变引起的单基因糖尿病患者的数据集中得到验证。我们还发现了HNF1A突变干细胞衍生β细胞代谢功能的表型，提供了HNF1A与II型和可能的I型糖尿病之间的潜在联系。最后，HNF1A的人类特异性靶点之一是一个长链非编码RNA，当在干细胞中删除时，模仿HNF1A突变体中看到的一些代谢表型。这些研究将有助于确定HNF1A在胰腺内分泌细胞中的发育和功能需求，并有可能直接影响罕见的单基因糖尿病以及I型和II型糖尿病的治疗。

Diabetes is a complex disorder with three main subtypes, type I, type II and monogenic. Type I is an autoimmune disorder characterized by loss of the insulin secreting beta cell. Type II is metabolic disorder characterized by poor insulin responsiveness and eventual beta cell exhaustion and loss. Monogenic forms are caused by mutations in a number of genes that are generally important in beta cell development and/or function. This proposal will focus on the study on HNF1A, a transcription factor that when mutated leads to the most common cause on monogenic diabetes. In addition, HNF1A has been linked to both type I and type II diabetes in GWAS studies. Mouse models of HNF1A deficiency are available, but unfortunately do not appropriately mimic the human disease. Therefore, use of a human model system is important. We will use the human pluripotent stem cell model to study the role of HNF1A in both endocrine cell development and function, utilizing CRISPR-Cas9 based genome engineering to generate isogenic mutant lines. Robust in vitro differentiation protocols exist and will be used to generate beta cells in the human stem cell model. In preliminary studies, we have uncovered human specific HNF1A targets that have been validated in a dataset from an individual with monogenic diabetes due to a HNF1A mutation. We have also discovered phenotypes in metabolic functions in HNF1A mutant stem cell derived beta cells, offering a potential link between the association of HNF1A with type II and possibly type I diabetes. Lastly, one of the human specific targets of HNF1A is a long non-coding RNA that when deleted in stem cells, mimics some of the metabolic phenotypes seen in the HNF1A mutants. These studies will help define the developmental and functional requirements of HNF1A in pancreatic endocrine cells and have the potential to directly impact the treatment of not only rare monogenic forms of diabetes but also type I and type II diabetes.