RNA localisation during the development of hyphae in the human pathogen Candida albicans

白色念珠菌是人类的主要病原体。它是女性阴道炎的病原体。此外，它还会在易受感染的病人中引起危及生命的血流感染，例如新生儿和某些重症监护病人，特别是那些正在接受癌症化疗、免疫抑制治疗或导尿术的病人。其生物学的一个显着特征是它能够以单细胞酵母或丝状形式生长，称为菌丝和伪菌丝。菌丝是由细长的细胞组成的链状结构，完全从菌丝顶端生长。这种转换生长形式的能力对毒力很重要。据认为，菌丝可以穿透表皮屏障，而酵母菌可以在血液中循环，从而在肾脏和心脏瓣膜建立致命的继发性感染。菌丝的生长仅限于它们的顶端；我们的研究重点是这种高度极化的生长形式的分子机制。如果我们了解了这些机制，就有可能通过找到阻止它们的方法来开发更好的抗真菌药物。然而，这是一个长期目标，不太可能作为这项研究的直接结果而实现。到目前为止，我们已经展示了一种名为Spitzenkörper的特殊结构位于尖端。据认为，结合在膜上的分泌囊泡含有新细胞壁和膜的原材料，被运输到Spitzenkörper，在那里积累，然后移动到细胞表面。因此，我们首先需要了解Spitzenkörper分泌囊泡聚集的机制。我们的研究重点是一种名为Sec2p的蛋白质，它与分泌囊泡有关，在它们向两极生长部位移动的过程中发挥着关键作用。这一建议是考虑到，当酵母细胞切换到菌丝生长时，细胞核保留在母酵母细胞中，母细胞与菌丝尖端可能有相当距离，在那里许多蛋白质需要聚集以促进极化生长。我们想知道，编码合成新蛋白质的信息的信使核糖核酸分子是否可能被特异性地从细胞核运输到菌丝顶端，在那里它们被翻译成新的蛋白质。为了验证这一想法，我们进行了一项初步实验，旨在检测Sec2p和mRNA分子之间的直接物理相互作用，并发现情况确实如此。我们现在希望确定附着在Sec2p上的单个mRNAs，并解决这样的问题：1)这些mRNAs编码的蛋白质在菌丝生长中扮演什么角色？2)编码这些蛋白质的mRNAs被运送到菌丝顶端是否重要？3)mRNAs是直接与Sec2p相互作用还是间接相互作用/即，mRNAs是否与包括Sec2p的分子复合体的另一种蛋白质相互作用？4)mRNAs是在分泌囊泡外部运输的吗？5)运输mRNAs的分子马达的性质和马达移动的轨迹的性质？这项拟议的研究利用了最新的基于基因组的技术以及最先进的显微镜技术。此外，这是白念珠菌细胞生物学和分子遗传学专家彼得·萨德贝里教授和研究细胞内信使核糖核酸运动的世界领先者斯图尔特·威尔逊博士共同提出的建议。

The fungus Candida albicans is a major human pathogen. It is the causative agent of vaginitis in women. In addition, it can cause life-threatening blood stream infections in vulnerable patients such as the new born and certain intensive care patients, especially those undergoing cancer chemotherapy, immunosuppressant therapy or catheterisation. A striking feature of its biology is its ability to grow as a unicellular yeast or in filamentous forms called hyphae and pseudohyphae. Hyphae are chains of long thin cells that grow exclusively from the hyphal tip. This ability to switch growth forms is important for virulence. It is thought that the hyphae can penetrate epidermal barriers while the yeast can circulate in the blood stream to establish lethal secondary infections in the kidneys and heart valves. Hyphal growth is restricted to their tip; our research focuses on the molecular mechanisms responsible for this highly polarised form of growth. If we understood these mechanisms, it might be possible to develop better anti-fungal drugs by finding ways to block them. However, this is a long term goal that is unlikely to be realised as a direct result of this research. So far, we have shown that a special structure called a Spitzenkörper is located at the tip. It is thought that membrane-bound secretory vesicles, which contain the raw materials for the new cell wall and membranes, are transported to the Spitzenkörper where they accumulate before being moving on to the cell surface. Thus we first need to understand the mechanisms responsible for the aggregation of secretory vesicles in the Spitzenkörper. Our research has focussed on a protein called Sec2p, which is associated with secretory vesicles and plays a key role in their movement to sites of polarised growth. This proposal is prompted by the consideration that as a yeast cell switches to hyphal growth the nucleus remains in the mother yeast cell which can be a considerable distance from the hyphal tip where many proteins need to aggregate to facilitate polarised growth. We wondered whether the mRNA molecules, which encode the information for the synthesis of new proteins, may be specifically transported from the nucleus, where they are transcribed from DNA, to the hyphal tip where they are translated into new proteins. To test this idea we carried out a preliminary experiment designed to detect a direct physical interaction between Sec2p and mRNA molecules, and found that this was indeed the case. We now wish to identify the individual mRNA molecules attached to Sec2p and resolve such questions as: 1) What is the role of the proteins encoded by these mRNA molecules in hyphal growth? 2) Is it important that mRNAs encoding these proteins are transported to the hyphal tip? 3) Do the mRNAs interact directly with Sec2p or is the interaction indirect / that is, do the mRNAs interact with another protein that is part of a molecular complex that includes Sec2p? 4) Are the mRNAs transported on the outside of secretory vesicles? 5) What is the nature of the molecular motors that transport the mRNA molecules and the nature of the tracks along which the motors move? The proposed research exploits the latest genome-based technologies as well as state-of-the-art microscopy. Moreover, this is a collaborative proposal between Professor Peter Sudbery an expert in Candida albicans cell biology and molecular genetics and Dr Stuart Wilson a world leader in the study of mRNA movement within cells.