Transcriptional Coregulators in Pancreas Development

描述（申请人提供）：分泌胰岛素的胰岛细胞的失败是1型和2型糖尿病的标志，导致本实验室强烈关注调节胰腺细胞发育和功能的分子机制。我们之前在酵母双杂交筛选中发现了Pdx1的伴侣Pcif1， Pdx1是一种同源结构域转录因子和人类糖尿病基因，在控制细胞质量的发育、成人细胞的功能和存活以及成人细胞补偿胰岛素抵抗增加的代谢需求的能力的转录层次中起关键作用。在之前的资助期内，我们确定Pcif1至少在一定程度上是通过其作为底物受体的作用，将Pdx1招募到cullin3基复合体中，用于泛素化和蛋白酶体降解，从而暗示Pcif1是调节Pdx1水平的靶标。事实上，Pcif1在细胞中的功能丧失会升高Pdx1蛋白并刺激重要Pdx1转录靶点的表达，包括胰岛素、MafA和Glut2。体内Pcif1的遗传破坏揭示了通过pdx1依赖和可能的pdx1独立机制在胚胎发育过程中调节成人细胞周转和功能以及内分泌谱系分配中的作用。我们假设Pcif1通过其调节关键转录调控因子的水平/功能的能力，对胚胎细胞形成和成人细胞周期进行关键调控。这一假设将在两个目标中得到验证：目标1。目的：探讨pci1如何调控成体细胞的周转率，而Aim 2如何调控成体细胞的周转率。确定pci1如何影响胚胎胰腺发育过程中细胞谱系的出现。从拟议的实验中获得的见解将为开发靶向治疗方法提供一个概念框架，这些治疗方法可以干扰pci1表达或其与细胞中关键底物的相互作用，从而影响细胞质量和细胞功能，并可能应用于涉及干细胞分化到细胞的方法，从其他成熟细胞类型的细胞重编程，促进糖尿病患者内源性β细胞团的功能和再生的途径。

DESCRIPTION (provided by applicant): Failure of insulin secreting pancreatic islet ¿ cells is a hallmark of both type1 and type 2 diabetes, leading to this laboratory's intense focus on molecular mechanisms regulating pancreatic ¿ cell development and function. We previously identified Pcif1 in a yeast two hybrid screen for partners of Pdx1, a homeodomain transcription factor and human diabetes gene pivotally positioned in the transcriptional hierarchy governing the development of ¿ cell mass, the function and survival of adult ¿ cells, and the ability of adult ¿ cells to compensate for increased metabolic demand imposed by insulin resistance. During the previous funding period we determined that Pcif1 acts at least in part through its role as a substrate adaptor to recruit Pdx1 into a cullin3-based complex for ubiquitination and proteasomal degradation, thereby implicating Pcif1 as a target for regulating Pdx1 levels. Indeed, Pcif1 loss of function in ¿ cells elevates Pdx1 protein and stimulates expression of important Pdx1 transcriptional targets, including insulin, MafA and Glut2. In vivo genetic disruption of Pcif1 reveals roles in regulating adult ¿ cell turnover and function and in endocrine lineage allocation during embryonic development, through Pdx1-dependent and, likely, Pdx1-independent mechanisms. We hypothesize that Pcif1 critically regulates embryonic ¿ cell formation and the adult ¿ cell cycle through its ability to modulate the levels/functions of key transcriptional regulators. This hypothesis will be tested in two aims: Aim 1. To determine how Pcif1 regulates the turnover of adult ¿ cells, and Aim 2. To determine how Pcif1 influences the emergence of the ¿ cell lineage during embryonic pancreas development. Insights gained from the proposed experiments will provide a conceptual scaffold for the development of targeted therapeutics that interfere with Pcif1 expression or its interactions with critical substrates in the ¿ cell to influence ¿ cell mass and ¿ cell function, with potential application to approaches involving stem cell differentiation to ¿ cells, cellular reprogramming of ¿ cells from other mature cell types, and for approaches to promote the function and regeneration of endogenous beta cell mass in patients with diabetes. PUBLIC HEALTH RELEVANCE: Failure of insulin secreting pancreatic islet ¿ cells is a hallmark of both type1 and type 2 diabetes, prompting intense effort to develop new sources of insulin- producing ¿ cells for replacement therapies and new ways to promote endogenous ¿ cell regeneration. The proposed studies will address important questions about the functions of the ubiquitin ligase substrate adaptor Pcif1 in ¿ cell biology. Insights gained will provide a conceptual scaffold for the development of targeted therapeutics that interfere with Pcif1 expression or its interactions with critical substrates in the ¿ cell to influence ¿ cell mass and ¿ cell function, with potential application to stem cell differentiation, cellular reprogramming and approaches to promote the function and regeneration of endogenous ¿ cell mass in patients with diabetes.