The role of OCRL1 in endocytic membrane traffic

我们体内的细胞是由被称为细胞器的内部膜包裹的隔间组成的，每个细胞器都有自己的身份和特定的功能。物质在细胞器之间以及某些细胞器与细胞外部之间通过一种称为膜运输的过程进行转移。主要的膜运输途径之一是内吞途径，物质通过吞噬在细胞表面的被膜包裹的小泡从细胞外部被带走。然后，内化的物质被分拣到各种下游隔间或回收回细胞表面。内吞途径对所有细胞都是基本重要的，当缺陷时会在人类中引起许多疾病。此外，致病病毒和细菌利用这一途径进入细胞造成感染。因此，重要的是我们要在分子水平上理解内吞途径的功能。这一提议集中在两种酶上，这两种酶定位于内吞途径中的细胞器，称为OCRL1和INPP5B。这些酶作用于一类被称为磷脂酰肌醇的脂类，这种脂类对控制细胞行为的各个方面，包括胞内膜运输非常重要。OCRL1基因突变会导致两种人类疾病，称为Lowe综合征和Dent-2病，其特征是大脑、眼睛和肾脏功能缺陷。OCRL1突变导致LOWE和Dent患者的病理变化的机制尚不清楚。我们已经确定了OCRL1和INPP5B的一个新的结合伙伴，称为IPIP27A，它对于内吞途径中的转运是必不可少的。这项提案的目的现在是确定IPIP27A如何与其互动伙伴一起发挥作用，以规范内吞贩运。我们的假设是OCRL1/INPP5B和IPIP27A参与内吞细胞器上膜载体的形成，以确保运输以适当控制的方式发生。这一过程是如何发生的，人们仍然知之甚少。我们计划使用实验方法的组合来剖析所涉及的机制。这将需要使用各种既定的方法来分析组织培养细胞的内吞运输，包括活细胞的实时成像。我们的工作将为了解内吞途径在所有细胞中的功能提供重要的知识，并揭示Lowe综合征、Dent病和其他由内吞转运缺陷引起的疾病的机制。

The cells in our body are made up of internal membrane-enclosed compartments called organelles, each with their own identity and specific functions. Material is transferred between the organelles and between certain organelles and the outside of the cell via a process termed membrane traffic. One of the major membrane traffic pathways is the endocytic pathway, where material is taken up from the cell exterior via engulfment in membrane-enclosed vesicles at the cell surface. The internalised material is then sorted to various downstream compartments or recycled back to the cell surface. The endocytic pathway is fundamentally important to all cells, and when defective gives rise to numerous diseases in humans. Moreover, this pathway is exploited by pathogenic viruses and bacteria to gain entry into cells to cause infection. It is therefore important we understand how the endocytic pathway functions at the molecular level. This proposal is focussed on two enzymes that are localised to organelles within the endocytic pathway, called OCRL1 and INPP5B. These enzymes act on a class of lipids called phosphoinositides that are important for controlling various aspects of cell behaviour including endocytic membrane traffic. Mutation of OCRL1 causes two diseases in humans, called Lowe syndrome and Dent-2 disease, which are characterised by defective brain, eye and kidney function. The mechanisms by which mutation of OCRL1 brings about the pathological changes seen in Lowe and Dent patients remain unclear. We have identified a new binding partner for OCRL1 and INPP5B called IPIP27A that is essential for trafficking within the endocytic pathway. The aim of this proposal is now to determine how IPIP27A functions together with its interaction partners to regulate endocytic trafficking. Our hypothesis is that OCRL1/INPP5B and IPIP27A participate in the formation of membrane carriers on endocytic organelles to ensure trafficking occurs in a properly controlled manner. How this process occurs remains poorly understood. We plan to use a combination of experimental approaches to dissect the mechanisms involved. This will entail analysis of endocytic trafficking in tissue culture cells using various established methods including imaging of living cells in real time. Our work will provide important knowledge as to how the endocytic pathway functions in all cells, as well as reveal the mechanisms underlying Lowe syndrome, Dent disease and other disorders caused by defects in endocytic trafficking.