The Role of the NPA-Binding, Plasma Membrane Aminopeptidase AtAPM1 in Cellular Trafficking and Auxin-Regulated Growth

哺乳动物的胰岛素反应性氨基肽酶(IRAP)和CD13(APN)等Gluzincin氨基肽酶是双功能蛋白，在细胞膜成分的转运中发挥重要作用。谷氨酸锌功能缺陷被认为是导致II型糖尿病和饮食中胆固醇摄取改变的原因之一。作为一种植物胶菌素，APM1似乎调节细胞机制，控制植物激素生长素的运动，这是建立和维持植物极性所必需的。与IRAP和APN一样，APM1是一种兼具酶功能和转运功能的双功能蛋白。APM1在多药耐药/P-糖蛋白的细胞转运中发挥作用，似乎还调节其他重要的发育机制。拟南芥apm1突变体表现出严重的发育缺陷，这种缺陷只能通过生长素处理部分改善。哺乳动物IRAP和APN对类固醇依赖的内吞循环的调节提示了APM1的一种机械功能。拟议研究的目标是确定APM1的作用机制，并确定它们对生长素运输的影响。提议的项目将是1。确定APM1在生长和发育中的作用位置和时间。本课程将研究APM1对生长素依赖和独立发育过程的调控。绘制APM1的膜拓扑图，并阐明APM1功能的机制。APM1对膜运输蛋白(包括那些运输生长素的蛋白)依赖于固醇的循环的调节将被研究。3.将APM1的功能指定为酶结构域和运输结构域，并确定负责运输活动的膜结构域。将确定特定的APM1蛋白结构域与膜转运蛋白的相互作用。拟议的项目具有创新性，因为它探索了以前未描述的植物中蛋白质运输和激素调节的机制。此外，这项拟议的研究可能会为谷氨酸锌蛋白在植物和动物中的功能提供新的见解。更广泛的影响：APM1功能的操纵可以用来改变植物的形态和生产力。因此，拟议的项目可能会增加农业和观赏作物的价值。此外，通过该项目加强对谷氨酸锌调节的了解，可以导致开发针对II型糖尿病、肥胖症和固醇代谢的新疗法。研究结果将在科学期刊和实验室网站上公布。重要的进展将在大众科学文献中公布。该项目还将培训至少一名博士生，掌握膜生物学、生理学、分子生物学和细胞生物学的综合方法。此外，该项目将每年招聘一名暑期本科生实习生，并对其进行相同技能的培训，以鼓励这些学生从事研究生研究教育和研究事业。实习生将从在基础生物学研究中为传统上代表性不足的人群提供服务的机构招聘。

Gluzincin aminopeptidases such as the mammalian Insulin Responsive Aminopeptidase (IRAP) and CD13 (APN) are dual function proteins that play essential roles in the cellular trafficking of membrane components. Defects in gluzincin function are thought to contribute to type II diabetes and altered uptake of dietary cholesterol. A plant gluzincin, APM1 appears to regulate cellular mechanisms that control the movement of the phytohormone auxin, which is required for the establishment and maintenance of plant polarity. Like IRAP and APN, APM1 is a bifunctional protein with both enzymatic and trafficking functions. APM1 functions in the cellular trafficking of Multiple Drug Resistance/ P-glycoproteins and appears to modulate other important developmental mechanisms as well. Arabidopsis apm1 mutants exhibit severe developmental defects that that can be only partially ameliorated by auxin treatment. A mechanistic function for APM1 is suggested by the regulation of sterol-dependent endocytotic cycling by mammalian IRAP and APN. The goal of the proposed research is to identify the mechanisms of APM1 action and determine their impact on auxin transport. The proposed project will1. Determine where and when APM1 functions in growth and development. APM1 regulation of auxin dependent and independent developmental processes will be examined.2. Map the membrane topology of APM1 and elucidate the mechanisms underlying APM1 function. APM1 regulation of sterol-dependent cycling of membrane transport proteins including those that transport auxin will be studied. 3. Assign APM1 functions to enzymatic and trafficking domains and identify the membrane domains responsible for trafficking activity. Interactions of specific APM1 protein domains with membrane transporters will be determined.The proposed project is innovative as it explores a previously undescribed mechanism of protein trafficking and hormonal regulation in plants. Further, the proposed research is likely to provide new insights into gluzincin protein function in both plants and animals. Broader Impacts: Manipulation of APM1 function can be used to alter plant form and productivity. As such, the proposed project can potentially increase the value of agricultural and ornamental crops. Further, enhanced understanding of gluzincin regulation derived from this project can lead to the development of new therapies for type II diabetes, obesity, and sterol metabolism. The results will be promulgated by publication in scientific journals and on the laboratory website. Important advances will be publicized in popular scientific literature.The project will also train at least one PhD student in integrated approaches to membrane biology, physiology, molecular biology, and cell biology. Further, the project will recruit and train one summer undergraduate intern each year in the same techniques in order to encourage those students to pursue a graduate research education and research career. Interns will be recruited from institutions serving traditionally under-represented populations in basic biological research.