Spatial relationships in pH signalling in a model filamentous fungus: roles of MVB pathway and plasma membrane components

生物体环境的一个至关重要的特征是酸度或碱度，即pH值。真菌调整它们分泌的酶的组合，它们合成的营养转运体，以及它们向其环境的pH值输出的产物（如毒素和抗生素）。真菌适应一系列环境pH值的能力对于它们感染植物和动物（包括人类）的能力以及它们产生黄曲霉毒素等毒素和青霉素和头孢菌素等抗生素的能力至关重要。我们的工作旨在确定环境碱度如何触发信号，最终导致dna结合蛋白的加工，使其能够“打开”适合碱性环境的真菌基因，并“关闭”适合酸性环境的真菌基因。先前的研究表明，有7种蛋白质专门参与其中一种是dna结合蛋白，另外6种参与传递pH信号。这六种信号蛋白包括位于细胞表面的两种，其中一种促进另一种可能是pH传感器的定位，一种蛋白质与一种表面蛋白质相互作用，两种蛋白质与细胞蛋白质分选机制的组成部分相互作用，其中一种蛋白质也与dna结合蛋白相互作用，一种蛋白质切割酶参与激活dna结合蛋白。此外，这种环境pH的信号传导涉及细胞内结构的一些成分，其其他工作是将某些蛋白质分类到细胞内的适当位置或走向破坏。在本项目中，我们将使用一种相对良性但与严重的机会致病菌烟曲霉、产生黄曲霉毒素的农业祸害黄曲霉和用于发酵生产清酒的丝状真菌米曲霉有关的模式丝状真菌——细粒曲霉（Aspergillus nidulans）。我们将确定哪些蛋白质分选成分参与环境pH的信号传递，并检查影响这些成分的突变的后果。在某些情况下，这些突变严重损害真菌的生长，但其他突变可以消除这种损害。我们将决定废除的机制。我们将确定pH信号的需求是否会导致蛋白质分选所需的那些成分的短缺，以及分选机制是否能够减弱pH信号。最后，我们将研究pH传感器在细胞表面的定位是如何由其他pH信号细胞表面蛋白辅助的。

A vitally important feature of the environment of an organism is the degree of acidity or alkalinity, i.e. the pH. Fungi tailor the combinations of the enzymes they secrete, the nutrient transporters they synthesise and the products (such as toxins and antibiotics) they export to the pH of their environment. The ability of fungi to adapt to a range of environmental pH is crucial to their ability to infect both plants and animals (including humans) and to their ability to produce toxins such as aflatoxin and antibiotics such as penicillin and cephalosporin. Our work is directed towards determining how a signal is triggered by environmental alkalinity, ultimately leading to the processing of a DNA-binding protein such that it is able to 'turn on' fungal genes whose action is appropriate to alkaline environments and 'turn off' those appropriate to acidic environments. Previous work has shown that seven proteins are exclusively involved of which one is the DNA-binding protein and the other six are involved in transmitting the pH signal. These six signalling proteins include two located at the cell surface, one of which facilitates localisation of the other which is probably the pH sensor, a protein interacting with one of the surface proteins, two proteins interacting with a component of the cellular protein-sorting machinery of which one also interacts with the DNA-binding protein and a protein-cutting enzyme involved in activating the DNA-binding protein. In addition, such signalling of environmental pH involves some components of intracellular structures whose other job is the sorting of certain proteins towards an appropriate location within the cell or towards destruction. In this project we will use the model filamentous fungus Aspergillus nidulans, a relatively benign organism but related to the serious opportunistic pathogen Aspergillus fumigatus, the aflatoxin-producing agricultural scourge Aspergillus flavus and the filamentous fungus used in the fermentation to produce sake Aspergillus oryzae. We shall determine which of the protein sorting components are involved in signalling environmental pH and examine the consequences of mutations affecting those components. In some cases these mutations seriously impair growth of the fungus but other mutations can abrogate that impairment. We shall determine the mechanism of abrogation. We shall determine whether the demands of pH signalling result in a shortage of those components also needed for protein sorting and whether the sorting machinery is able to attenuate pH signalling. Finally we will investigate how the localisation of the pH sensor at the cell surface is assisted by the other pH signalling cell surface protein.