Role of PDX1 in insulin biosynthesis and ER homeostasis

描述（由申请人提供）： 近几十年来，美国经历了一个有增无减的个人数量上升， 被诊断患有糖尿病;因此，目前有2300万人患有2型糖尿病。的 2型糖尿病的发病机制涉及胰岛素抵抗和胰岛素抵抗失败之间的复杂相互作用。 胰腺细胞代偿细胞补偿通过改善细胞功能和细胞质量扩张发生，这两者都由关键的胰腺转录因子PDXI调节。初步数据显示，PDXI缺乏通过增加对内质网（ER）应激的敏感性，在饮食诱导的胰岛素抵抗期间增加细胞凋亡（M。Sachdeva，未发表的数据，2008年）。一个细胞的ER每分钟处理一百万个胰岛素分子，ER应激与小鼠和人类的2型糖尿病有关。高通量基因表达和启动子占用分析表明，PDXI调节参与维持ER稳态的几个基因，包括EROlip和GRP 170。假设EROlip和GRP 170的PDXI调节对于胰岛素生物合成和细胞存活是重要的。这一假设将在以下具体目标进行检验：I。确定DX 1在胰岛素生物合成中的作用，II。确定PDXI在调节ER稳态和p细胞存活中的作用。将在具有shRNA沉默的EROlip和GRP 170的小鼠胰岛素瘤细胞中研究胰岛素分泌。将分析EROlip在胰岛素二硫键形成中的作用和GRP 170作为细胞伴侣的功能。为了确定EROlip和GRP 170缺乏对ER稳态的影响，将测量ER应激标志物如BiP和CHOP，并使用膜联蛋白V和TUNEL染色来定量细胞凋亡。EROlip和GRP 170的过表达可以部分挽救PDXI缺陷表型，表明它们是PDXI对胰岛素分泌和细胞存活的影响的重要介质。 相关性：细胞不能充分分泌胰岛素和这些产生胰岛素的细胞死亡导致糖尿病。研究PDXI调节胰岛素分泌和细胞存活的机制可能会导致治疗2型糖尿病的目标。

DESCRIPTION (provided by applicant): In recent decades, the United States has experienced an unabated rise in the number of individuals diagnosed with diabetes; as a result 23 million people are currently living with type 2 diabetes. The pathogenesis of type 2 diabetes involves a complex interplay between insulin resistance and failure of pancreatic ¿ cell compensation. ¿ cell compensation occurs through improved ¿ cell function and ¿ cell mass expansion, which are both regulated by the critical pancreatic transcription factor PDXI. Preliminary data show that PDXI deficiency increases ¿ cell apoptosis during diet induced insulin resistance by increasing susceptibility to endoplasmic reticulum (ER) stress (M. Sachdeva, unpublished data, 2008). The ER of a ¿ cell processes one million insulin molecules per minute, and ER stress has been associated with type 2 diabetes in mice and humans. High throughput gene expression and promoter occupancy analyses suggest that PDXI regulates several genes involved in maintaining ER homeostasis, including EROlip and GRP170. PDXI regulation of EROlip and GRP170 is hypothesized to be important for insulin biosynthesis and ¿ cell survival. This hypothesis will be tested in the following specific aims: I. Determine the role of DX1 in insulin biosynthesis, II. Determine the role of PDXI in the regulation of ER homeostasis and p cell survival. Insulin secretion will be studied in mouse insulinoma cells with shRNA silenced EROlip and GRP170. The role of EROlip in insulin disulfide bond formation and the function of GRP170 as a ¿ cell chaperone will be analyzed. To determine the effects of EROlip and GRP170 deficiency on ER homeostasis, ER stress markers such as BiP and CHOP will be measured, and Annexin V and TUNEL staining will be used to quantify ¿ cell apoptosis. Overexpression of EROlip and GRP170 may partially rescue the PDXI deficiency phenotype suggesting that they are important mediators of the effects of PDXI on insulin secretion and ¿ cell survival. Relevance: Failure of ¿ cells to sufficiently secrete insulin and death of these insulin-producing cells results in diabetes. Studying the mechanism for PDXI regulation of insulin secretion and ¿ cell survival may lead to targets for treatment of type 2 diabetes.