The Role of Clec16a in the Pancreatic Islet

本计划书详细介绍了Scott Soleimanpour医学博士的研究和培训计划，他将作为一名医学科学家，专注于深入了解调节胰腺细胞复制和存活的因素，最终目标是改善1型糖尿病的治疗。拟议的K08奖的职业发展部分包括高级教学课程和正式的研究指导，在主要导师的监督下，Doris a . stoffer，医学博士，宾夕法尼亚大学医学院的医学副教授，以及一个多学科指导委员会。宾夕法尼亚大学医学院提供了一个研究丰富的环境，可以开放访问各种各样的专家教师和资源，这将有利于Soleimanpour博士的科学和专业发展。本次K08申请的研究将重点关注1型糖尿病易感基因、细胞必需转录因子Pdx1的新靶点Clec16a （c -凝集素结构域家族16，成员A）的细胞特异性作用。果蝇Clec16a同源物Ema是一种内体蛋白，通过与Vps-C HOPS复合物的直接相互作用对内体成熟至关重要，该复合物调节内体和自噬体成熟的关键转变。Ema功能的丧失导致生长因子信号的增加，这是由于配体结合受体的清除减少，从而导致突触末端过度生长。虽然Clec16a被认为在免疫细胞中表达受限，但我们的初步证据表明，Clec16a在小鼠和人类胰岛中高度表达。此外，功能缺失实验表明，Clec16a调节晚期核内体积累、细胞复制、胰岛素分泌和线粒体呼吸。因此，我们假设Clec16a在体内调节细胞内体途径和维持细胞质量和功能。1 .验证Clec16a对细胞质量和葡萄糖稳态维持至关重要的假设。该目标将通过使用条件Clec16a loxP等位基因的体内细胞特异性功能丧失实验来确定Clec16a的作用。实验将集中在葡萄糖耐量和胰岛素分泌的测定，以及与细胞增殖和/或存活变化相关的细胞质量的形态学分析。具体目标二。为了验证Pdx1靶点Clec16a调节核内体、自噬体和线粒体功能的基本方面的假设。实验将确定Clec16a在核内体和自噬体成熟、核内体运输、线粒体生物发生和钙通量中的作用。此外，Pdx1在调控Clec16a和¿-细胞内体途径中的上游调控作用将被确定。通过这些具体目标，我们将建立一个新的Pdx1靶点在调节细胞质量和功能中的作用和功能机制。我们还将确定内体途径中一个靶点对细胞功能的重要性，包括胰岛素分泌、复制/存活、线粒体功能和自噬调节，这可能为未来的研究开辟一个新的领域。

DESCRIPTION (provided by applicant: This proposal details the research and training plan for Scott Soleimanpour, M.D. to develop his academic career as a physician-scientist focused on the thorough understanding of factors which regulate pancreatic ¿-cell replication and survival, with the eventual goal of improving treatment of type 1 diabetes. The career development portion of the proposed K08 award includes an advanced didactic curriculum and formal research mentoring under the supervision of the primary mentor, Doris A. Stoffers, M.D., Ph.D., Associate Professor of Medicine at the University of Pennsylvania School of Medicine, and a multi-disciplinary mentorship committee. The University of Pennsylvania School of Medicine offers a research-rich environment with open access to a wide assortment of expert faculty and resources that will be beneficial in the scientific and professional development of Dr. Soleimanpour. The research proposed in this K08 application will focus on the ¿-cell specific role of Clec16a (C-lectin domain family 16, member A), a type 1 diabetes susceptibility gene and novel target of the essential ¿-cell transcription factor Pdx1. The Drosophila orthologue of Clec16a, Ema, is an endosomal protein vital to endosomal maturation through direct interaction with the Vps-C HOPS complex, which regulates key transitions in endosome and autophagosome maturation. Loss of Ema function leads to increased growth factor signaling due to decreased clearance of ligand bound receptors with resultant synaptic terminal overgrowth. While Clec16a has been suggested to have restricted expression to immune cells, our preliminary evidence indicates that Clec16a is highly expressed in mouse and human islets. Further, loss of function experiments suggest that Clec16a regulates late endosome accumulation, cell replication, insulin secretion, and mitochondrial respiration in ¿-cell lines. Therefore, we hypothesize that Clec16a regulates the ¿ -cell endosomal pathway and the maintenance of ¿-cell mass and function in vivo. Specific Aim I. To test the hypothesis that Clec16a is critical to ¿-cell mass and maintenance of glucose homeostasis. This aim will determine the role of Clec16a via ¿-cell specific loss-of-function experiments in vivo using a conditional Clec16a loxP allele. Experiments will focus on determinations of glucose tolerance and insulin secretion, as well as morphologic analysis of ¿-cell mass related to changes in ¿ -cell proliferation and/or survival. Specific Aim II. To test the hypothesis that the Pdx1 target Clec16a regulates essential aspects of endosome, autophagosome, and mitochondrial function. Experiments will determine the role of Clec16a in endosomal and autophagosome maturation, endosomal trafficking, and mitochondrial biogenesis and calcium flux. In addition, the upstream regulatory role of Pdx1 in regulation of Clec16a and the ¿ -cell endosomal pathway will be determined. Through these specific aims, we will establish the role and functional mechanism for a novel Pdx1 target in the regulation of ¿ -cell mass and function. We will also determine the importance of a target in the endosomal pathway to ¿ -cell function, including insulin secretion, replication/survival, mitochondrial function, and regulation of autophagy, that may open a new areas for future research. PUBLIC HEALTH RELEVANCE: This proposal will focus on discovering the role of Clec16a (KIAA0350), a gene recently identified to be associated with type 1 diabetes in humans. We know very little of the function of this gene in the insulin producing ¿ -cells of the pancreas. We are unsure if loss of this gene leads the ¿ -cells to die or not produce insulin properly. Therefore, our mission is to critically study this gene in order to understand how this gene affects the ¿-cell with hopes that our findings will direct future approaches to treat diabetes.