The function of the ESCRT machinery in the maintenance of the septate junction of Epithelia in Drosophila

ESCRT 机制在维持果蝇上皮细胞隔膜连接中的功能

• 批准号: 420088258
• 负责人: Professor Dr. Thomas Klein
• 金额: --
• 依托单位: Institut für Genetik
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2019
• 资助国家: 德国
• 起止时间: 2018-12-31 至 2021-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/420088258?language=en
• 关键词: function; ESCRT; machinery; maintenance; septate

Epithelia cover most surfaces of our body and regulate its communication and substrate exchange with the environment. They are the origin of the majority of solid cancers in humans. The basis for their functionality is a strictly polarized organisation along their apico-basal axis, which is lost during cancerogenesis. In the last decade it has been found that the function of the ESCRT machinery is essential for the integrity of the wing disc epithelium of Drosophila, a major model system for studying epithelia. How the ESCRT machinery, which is involved in the formation of intraluminal vesicles (ILVs) during endosomal trafficking of transmembrane proteins (TMPs) to the lysosome, maintains epithelial polarity and controls cell proliferation is not well understood. We have developed an RNAi-based system where we can generate weak loss of function situations of the ESCRT core component Shrub. We found that the septate junction is the most sensitive epithelial structure towards loss of ESCRT function. Moreover, Shrub/ESCRT appears to be involved in the trafficking of newly synthesised Megatrachea (Mega) to the SJ. This route also requires an unidentified variant of the Vps-Retromer, which is normally involved in endosomal recycling. We therefore discovered a new trafficking route of SJ components to the apical membrane, which is important for the maintenance of SJ in an already established epithelium. Our data suggest that ESCRT regulates Retromer dependent trafficking of Mega from the basal to its apical destination, thus mediating a transcytosis like event. In this application, we will investigate how Mega and other SJ components are transported to the SJ and what the role of the ESCRT machinery is in this process. In addition, we found that the expression of the EMT (epithelial mesenchymal transformation) factor Snail is induced by the depletion of Shrub. The activation of this factor has been shown to induce neoplastic transformation and could therefore explain the induction of neoplastic growth seen upon depletion of Shrub function. We will therefore find out how Snail is induced and how it contributes to the neoplastic transformation induced by Shrub depletion.For our analysis, we will use a combination of genetic, biochemical and molecular tools, as well as electron-, confocal live imaging and super-resolution microscopy. Our results will provide a deeper understanding of a novel function of the ESCRT machinery as a key organizer of epithelial integrity and endosomal trafficking. Moreover, they will also clarify its role during neoplastic transformations of epithelia and therefore contribute to the understanding of EMT, which is also an important event during cancerogenesis.

上皮细胞覆盖我们身体的大部分表面，并调节其与环境的通讯和基质交换。它们是人类大多数实体癌的起源。其功能的基础是沿着其顶端-基底轴的严格极化组织，其在癌发生期间丢失。在过去的十年中，人们已经发现，ESCRT机制的功能是必不可少的果蝇翅盘上皮细胞，研究上皮细胞的主要模型系统的完整性。在跨膜蛋白（TMP）的内体运输到溶酶体期间，ESCRT机制如何参与管腔内囊泡（ILV）的形成，并维持上皮极性和控制细胞增殖还不清楚。我们已经开发了一种基于RNAi的系统，在该系统中，我们可以产生ESCRT核心组件Shrub的弱功能丧失情况。我们发现隔膜连接是对ESCRT功能丧失最敏感的上皮结构。此外，Shrub/ESCRT似乎参与了向SJ贩运新合成的Megalehea（Mega）。该途径还需要Vps-Retromer的未鉴定变体，其通常参与内体再循环。因此，我们发现了一个新的运输路线SJ组件的顶端膜，这是重要的SJ在已经建立的上皮细胞的维护。我们的数据表明，ESCRT调节Retromer依赖的交通的巨型从基底到其顶端的目的地，从而介导转胞吞样事件。在这个应用程序中，我们将研究如何兆和其他SJ组件被运送到SJ和什么样的作用的ESCRT机器在这个过程中。此外，我们发现EMT（上皮间质转化）因子Snail的表达是由Shrub的耗竭诱导的。该因子的激活已被证明可诱导肿瘤转化，因此可以解释在Shrub功能耗尽后观察到的肿瘤生长诱导。因此，我们将找出蜗牛是如何被诱导的，以及它如何有助于灌木枯竭诱导的肿瘤转化。对于我们的分析，我们将使用遗传，生化和分子工具的组合，以及电子，共聚焦实时成像和超分辨率显微镜。我们的研究结果将提供一个新的功能的ESCRT机器作为上皮完整性和内体运输的关键组织者的更深入的了解。此外，他们还将阐明其在上皮细胞肿瘤转化过程中的作用，因此有助于理解EMT，EMT也是癌症发生过程中的重要事件。