PTPN2 mutations affect islet beta cell susceptibility in T1D

1型糖尿病(T1D)的特征是个体自身的免疫系统破坏胰岛β细胞。尽管T1D主要被视为一种免疫系统疾病，但越来越多的证据表明，产生胰岛素的β细胞可能积极地促进了它们的破坏。此外，最近的全基因组关联研究表明，尽管人类白细胞抗原(HLA)复合体决定了T1D的显著遗传易感性，但非HLA基因座上的许多单核苷酸多态(SNPs)也与T1D有关(Floyel，2015#2)。重要的是，这些非人类白细胞抗原变异体中的许多存在于免疫细胞系以及靶向β细胞中表达的基因中。了解这些微小的基因变异如何影响胰岛b细胞的功能，将使我们能够开发出有效防止T1D进展的新疗法。在这个方案中，我们将使用转基因小鼠和人类b细胞模型来探索在T1D和/或T1D模拟条件下，T1D风险等位基因蛋白酪氨酸磷酸酶N2(也称为T细胞蛋白酪氨酸磷酸酶)的突变如何影响胰岛β细胞的功能。在小鼠和人类中，PTPn2在包括T细胞和胰岛β细胞在内的多种组织中广泛表达，然而，PTPn2的突变主要是在T细胞背景下进行的研究。在小鼠中，PTPN2被证明通过作为细胞因子反应下游的JAK/STAT信号的负调节因子来调节T细胞中的炎症信号。然而，有证据表明，PTPN2介导的自身免疫不能完全用T细胞功能障碍来解释。与全局基因敲除小鼠相比，PTPN2的T细胞特异性失活导致的自身免疫表型不那么严重。事实上，在胰腺中，PTPN2的结构性失活损害了高脂肪饮食小鼠的葡萄糖刺激的胰岛素分泌[xi，2015#20}。PTPn2在β细胞中的功能尚未在T1D的背景下进行检测；然而，在促炎细胞因子或双链病毒RNA处理下，PTPn2在小鼠和人β细胞中上调，这再次表明PTPn2在胰岛中具有额外的作用。为了分析PTPn2突变对T1D的β细胞特异性贡献，我们产生了β细胞特异性的PTPn2基因敲除(PTPn2-BKO)。我们的初步数据表明，在模拟T1D应激条件下，PTPN2-BKO胰岛的代谢途径发生了变化。一直以来，我们都发现β细胞中的重要糖酵解酶--丙酮酸激酶M2(PKM2)是PTPN2的直接靶标。此外，我们还生成了PTPN2缺失的hPSCs，并收集了携带PTPN2突变的T1D个体的组织样本。在这个提案中，我们将检验这样的假设，即β细胞中PTPn2功能的中断通过损害β细胞的代谢功能和生存来促进T1D的发展。在SA1中，我们将确定在基础和自身免疫介导的条件下，与PTPN2缺失相关的β细胞缺陷。在SA2中，我们将确定在基础和自身免疫介导的条件下，PTPN2调节的分子通路。

Type 1 diabetes (T1D) is characterized by the destruction of pancreatic beta cells by an individual’s own immune system. Although T1D is primarily viewed as a disease of the immune system, there is mounting evidence to suggest that insulin-producing β cells may actively contribute to their destruction. Furthermore, recent genome-wide association studies (GWAS) have suggested that although significant genetic predisposition to T1D is conferred by the human leukocyte antigen (HLA) complex, many single nucleotide polymorphisms (SNPs) in non-HLA loci also contribute to the disease {Floyel, 2015 #2}. Importantly, many of these non-HLA variants occur in genes expressed in both immune cell lineages as well as in the targeted β cells. Knowledge of how these minor gene variants affect function of islet b cells will allow us to develop novel therapies that could be effective in preventing progression of T1D. In this proposal, we will use transgenic mice and human b cell models to explore how mutations in the T1D risk allele Protein tyrosine phosphatase N2 (PTPN2), also known as T cell protein tyrosine phosphatase (TC-PTP), affect islet β cell function in the context of T1D and/or T1D mimicking conditions. In mice and humans, PTPN2 is broadly expressed in several tissues, including T cells and islet β cells, however mutations of PTPN2 have predominantly been studied in the T cell context. In mice, PTPN2 has been show to modulate inflammatory signaling in T cells by acting as a negative regulator of JAK/STAT signaling, downstream of a cytokine response. However, there is evidence that PTPN2-mediated autoimmunity cannot be fully explained by T cell dysfunction. T-cell specific inactivation of PTPN2 caused less severe autoimmune phenotypes than observed in global knockout mice. Indeed, in the pancreas, constitutive inactivation of PTPN2 impaired glucose stimulated insulin secretion in mice fed a high fat diet {Xi, 2015 #20}. The function of PTPN2 in β cells has not yet been examined in the context of T1D; however, PTPN2 is upregulated in mouse and human β cells upon treatment with proinflammatory cytokines or double stranded viral RNA, again suggesting an additional role for PTPN2 in islets. To parse out the β cell-specific contribution of PTPN2 mutations to T1D, we have generated a β cell-specific knockout of Ptpn2 (PTPN2-bKO). Our preliminary data indicate that metabolic pathways are altered in PTPN2-bKO islets under T1D mimicking stress conditions. Consistently, we have identified pyruvate kinase M2 (PKM2), an important glycolytic enzyme in the β cell to be a direct target of PTPN2. In addition, we have generated hPSCs deleted for PTPN2 and collected tissue samples from T1D individuals carrying mutations in PTPN2. In this proposal, we will test the hypothesis that disrupted function of PTPN2 in the β cell promotes the development of T1D by compromising β cell metabolic function and survival. In SA1 we will determine the beta cell defects associated with deletion of PTPN2 in basal and autoimmune-mediated conditions. In SA2 we will determine the molecular pathways regulated by PTPN2 in basal and autoimmune-mediated conditions.