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.

上皮涵盖了我们身体的大多数表面，并调节其与环境的沟通和底物交换。它们是人类大多数固体癌的起源。其功能性的基础是沿其Apico-Basal轴的严格极化组织，在癌发生过程中丢失。在过去的十年中，已经发现，ESCRT机械的功能对于果蝇的翼盘上皮的完整性至关重要，果蝇是研究上皮的主要模型系统。在跨膜蛋白（TMPS）向溶酶体的内体运输过程中，如何参与腔内囊泡（ILV）形成的ESCRT机械如何保持上皮极性并控制细胞增殖。我们已经开发了一个基于RNAi的系统，可以在其中产生ESCRT核心成分灌木功能情况弱损失。我们发现，分隔连接是损失ESCRT功能的最敏感的上皮结构。此外，灌木/escrt似乎参与了新合成的巨型杂质（Mega）贩运到SJ。该路线还需要VPS-ReTromer的未识别变体，该变体通常参与内体回收。因此，我们发现了SJ组件到根尖膜的新贩运途径，这对于在已经建立的上皮中维持SJ很重要。我们的数据表明，ESCRT调节大型从基础到其顶端目的地的逆转录依赖性运输，从而介导类似事件的跨经细胞增多症。在此应用程序中，我们将调查如何将巨型和其他SJ组件传输到SJ，以及ESCRT机械在此过程中的作用。此外，我们发现EMT（上皮间充质转化）因子蜗牛的表达是由灌木的耗竭诱导的。该因子的激活已被证明会诱导肿瘤转化，因此可以解释灌木功能消耗后观察到的肿瘤生长的诱导。因此，我们将找出蜗牛是如何诱导的，以及它如何促进灌木消耗引起的肿瘤转化。对于我们的分析，我们将使用遗传，生化和分子工具以及电子，共核活性成像和超分辨率显微镜的组合。我们的研究结果将对ESCRT机械作为上皮完整性和内体贩运的关键组织者的新功能有更深入的了解。此外，他们还将阐明其在上皮症的肿瘤转化中的作用，因此有助于对EMT的理解，这也是癌发生过程中的重要事件。