Fungal Pheromone Receptors in Host-Pathogen Interactions

g蛋白偶联受体（gpcr）形成了一个细胞表面受体的超家族，在真核生物分类群中发现，例如，仅在人类中就有800个独特的gpcr。真菌的Ste2p和Ste3p gpcr介导交配反应，通过信息素对受体的刺激导致有充分记录的信号转导事件、细胞周期阻滞和细胞融合。然而，最近的一些发现表明，这些信息素受体的相关性和相关机制可能比这更广泛。一方面，最近的一份报告表明，Ste2p参与了致病真菌对宿主的过氧化物酶介导的化学吸引。另一方面，发现了“偏倚信号”，即多个不同的配体可以刺激单个gpcr，每个配体刺激不同的G亚基和抑制因子。我们自己最近的工作已经获得了证据，强调了酵母Ste2p在经典细胞周期阻滞与下游细胞壁降解/投射形成中的偏倚功能的可能性，以及阻滞在介导这些后一种反应中的可能作用。****这些发现提出了一个假设，即宿主-病原体化学吸引的Ste2p相关机制在病原真菌中是保守的，并且刺激反应的配体的性质是一种过氧化物酶修饰的细胞壁成分，选择性地刺激抑制蛋白相关的偏倚信号通路。我们将通过真菌遗传实验、半靶向代谢组学以及结合位点定向诱变、功能分析和结构研究的长期经典重组分析来验证这一假设。该研究项目将培养遗传学、细胞生物学和生物化学（包括代谢组学和计算/结构生物学）领域的3名硕士和2名博士研究生。****真菌感染是一个广谱问题，影响人类和牲畜的健康以及大多数农作物的生产力。本提案的目标是谷物赤霉病（Fusarium Head Blight， FHB）的病原体——镰刀菌（Fusarium graminearum），它包括有害真菌毒素的沉积。每年仅在加拿大，食品和卫生局就造成了1亿美元的经济损失，并在更广泛的范围内威胁着世界粮食供应。常用的抗真菌药物一般都是利用真菌细胞膜的可塑性起作用的。然而，耐药性正在发展，目前特异性的缺乏影响了环境中的其他生物。开发具有替代作用模式的新的更特异的抗真菌药是当务之急。为了实现这一点，我们必须对真菌感染的基础有一个完整的了解。*********

G-protein Coupled Receptors (GPCRs) form a superfamily of cell surface receptors that are found across eukaryotic taxa, with, for example >800 unique GPCRs in humans alone. The fungal Ste2p and Ste3p GPCRs mediate mating responses, with stimulation of the receptors by pheromones leading to well-documented signal transduction events, cell-cycle arrest and cellular fusion. However a number of recent findings suggest the relevance and associated mechanisms of these pheromone receptors may be much broader than this. On the one hand, a recent report has implicated Ste2p in the peroxidase-mediated chemo-attraction of pathogenic fungi to hosts. On the other hand, is the discovery of biased-signalling' where individual GPCRs can be stimulated by multiple different ligands, each stimulating different G subunits and arrestins. Our own recent work has yielded evidence highlighting the possibility of biased' functionalities for yeast Ste2p in classical cell cycle arrest versus downstream cell wall degradation/projection formation, and a possible role for arrestins in mediating these latter responses.****Together these findings raise the hypothesis that the Ste2p associated mechanism of host-pathogen chemo-attraction is conserved across pathogenic fungi, and that the nature of the ligand stimulating the response is a peroxidase-modified cell wall component selectively stimulating an arrestin-associated biased-signaling pathway. We will test this hypothesis through a combination of fungal genetic experiments, semi-targeted metabolomics, and in the longer-term, classical recombinant analyses combining site directed mutagenesis with functional assays and structural investigations. This program of research will enable the training of 3 MSc and 2 PhD graduate students in the fields of genetics, cell biology and biochemistry (incl. metabolomics and computational/structural biology).****Fungal infection is a broad-spectrum problem, affecting the health of humans and livestock as well as the productivity of most agricultural crops. Targeted in this proposal is Fusarium graminearum, the causative agent of Fusarium Head Blight (FHB) in cereal crops, which includes deposition of hazardous mycotoxins. FHB causes $ 100's of millions in economic losses annually in Canada alone, and threatens world food supplies more broadly. Commonly used antifungals generally all work by targeting the plasticity of the fungal cell membrane. However resistance is developing, and the current lack of specificity impacts other organisms in the environment. It is imperative that new more specific antifungals, with alternate modes of action, are developed. To enable this, it is essential that we have a complete understanding of the basis of fungal infection.*********