Protein targeting to endosomes/lysosomes: function of Septins in AP-3-dependent transport

靶向内体/溶酶体的蛋白质：Septins 在 AP-3 依赖性转运中的功能

• 批准号: 90048439
• 负责人: Professor Dr. Bernard Hoflack
• 金额: --
• 依托单位: Biotechnologisches Zentrum (Biotec)
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2008
• 资助国家: 德国
• 起止时间: 2007-12-31 至 2012-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/90048439?language=en
• 关键词: Protein; targeting; endosomes; lysosomes; function

Clathrin-coats are involved in various aspects of lysosome biogenesis. Clathrin associates with hetero-tetrameric adaptor proteins (APs). While AP-2 functions in endocytosis, the ubiquitously expressed AP-1 A, AP-3 and AP-4 are required for the transport of transmembrane proteins from the secretory pathway to the endosomal/lysosomal system. Using a liposome-based in vitro systems that recapitulate the fidelity of protein sorting in vivo, we recently identified the protein networks supporting AP-1 and AP-3 sorting functions, which collectively highlight the functional importance of =80 proteins. In this proposal, we will focus on the functional characterization of the protein networks supporting AP-3 sorting function in targeting of lysosomal membrane associated proteins. In particular, we will investigate the function of several septin family members and their regulator Borg4 in this process. Our working hypothesis is that septins, by interacting with proteins of the AP-3 sorting machinery, facilitate the binding of AP-3-coated membranes onto microtubules and facilitate microtubule-dependent transport from early to late endosomes. To test this hypothesis, it will be necessary 1) to colocalize septins and AP-3, 2) to establish the functional importance of septins in membrane traffic along the endocytic pathway, in particular during the early to late endosome transition, 3) to test whether septins are found along specific microtubule tracks, 4) to establish the molecular links between septins and their effector Borg4 with other proteins belonging to the AP-3 network, and 5) to understand how septin function is regulated by Borg4, and the cdc42 GTPase and by phosphorylation.

网格蛋白被膜参与溶酶体生物发生的各个方面。网格蛋白与异源四聚体衔接蛋白（AP）缔合。虽然AP-2在内吞作用中起作用，但普遍表达的AP-1A、AP-3和AP-4是跨膜蛋白从分泌途径转运到内体/溶酶体系统所需的。使用脂质体为基础的体外系统，概括了蛋白质分选在体内的保真度，我们最近确定的蛋白质网络支持AP-1和AP-3的分选功能，共同突出的功能重要性=80蛋白质。在这个提议中，我们将集中在支持AP-3分选功能的蛋白质网络的功能特性，在靶向溶酶体膜相关蛋白。特别是，我们将研究几个septin家族成员和他们的调节Borg 4在这个过程中的功能。我们的工作假设是，septins，通过与AP-3分选机制的蛋白质相互作用，促进AP-3包被的膜结合到微管上，并促进微管依赖性运输从早期到晚期内体。为了检验这一假设，有必要1）共定位隔蛋白和AP-3，2）确定隔蛋白在沿着内吞途径的膜运输中的功能重要性，特别是在早期到晚期的内体转换期间，3）检验是否发现隔蛋白沿着沿着特定的微管轨道，4）建立septins及其效应物Borg 4与属于AP-3网络的其他蛋白质之间的分子联系，以及5）了解septin功能如何受Borg 4调节，和cdc 42 GT3的表达以及磷酸化。