Phenotypic Plasticity and Inter-Species Communication in Candida Albicans

描述(申请人提供)：致病酵母菌，白色念珠菌是人类菌群的主要组成部分，具有可塑性，使其能够作为共生有机体和威胁生命的病原体茁壮成长。90%的HIV感染者患上口咽念珠菌病(OPC)，口腔念珠菌感染也影响5%的新生儿和10%的老年患者(2；3)。这种真菌变色龙通过对外部提示的反应来调节形态、表型和菌落结构的变化，这些变化赋予白色念珠菌增加毒力、增强定植、允许交流和抵抗抗真菌药物的特征。这项建议中包含的实验解决了这种可塑性的关键特征，这些特征对白色念珠菌的致病性至关重要：环境信号和物种间交流如何引发表型转换和信息素信号通路，这两个过程都直接与生物膜的形成有关。我的初步工作表明，在氧化应激和常用的抗真菌药物呋喃胞嘧啶的存在下，调节生物膜形成和交配的白色不透明开关会增加。鉴于这些结果，在我的第一个目标中，我将确定白色念珠菌对氧化应激、亚硝化应激和抗真菌药物暴露的反应转化率。这些因素是临床环境中白色念珠菌面临的共同挑战，因此，将对几种临床分离株进行分析。我还将确定它们增加表型转换的反应途径。我假设硫氧还蛋白途径和RAD53途径参与其中，因此将使用TRX1、TRR1、Cap1和RAD53基因缺失和过表达菌株进行测试。我的第二个目标将解决白色念珠菌与人类口腔微生物组的真菌和细菌成员之间的物种间事件。来自我的实验室的结果表明，来自其他念珠菌的信息素信号诱导白色念珠菌形成生物膜，我将在这个目标中进一步研究(14)。首先，我将使用我构建的一组白色念珠菌菌株来确定假丝酵母菌之间的信号是否在生理上相关的信息素浓度下发生，这些菌株分泌这些其他物种本身的信息素，而不是自己的信息素。我预计，我的实验将表明，交配和生物膜的形成都是通过几个念珠菌物种的信息素对信号做出反应，这意味着真菌间的信号在发病中。此外，我将直接与已知细菌共培养白色念珠菌 口腔微生物组的成员，包括戈登链球菌，它分泌高水平的过氧化氢，一种已知的应激源和转换诱导剂(43)。我的第二个目标的结果将得到我的第一个目标的支持。了解这种白色不透明的开关和物种间信号事件将有助于开发新的治疗方法，以防止与这种真菌病原体相关的往往使人虚弱，有时甚至是致命的感染。

DESCRIPTION (provided by applicant): The pathogenic yeast, Candida albicans is a major constituent of the human flora with a plasticity that allows it to thrive both as a commensal organism and as a life-threatening pathogen. 90% of HIV-infected individuals develop oropharyngeal candidiasis (OPC), and oral Candida infections also affect 5% of newborns and 10% of elderly patients (2; 3). This fungal chameleon mediates changes in morphology, phenotype, and colony structure by responding to external cues, and these alterations endow C. albicans with traits that increase its virulence, enhance colonization, allow communication and resist anti-fungal drugs. The experiments contained in this proposal address key features of this plasticity that are paramount to C. albicans pathogenicity: how environmental signals and inter-species communication instigate phenotypic switching and pheromone signaling pathways, both of which are processes directly implicated in biofilm formation. My preliminary work shows that the white-opaque switch, which regulates both biofilm formation and mating, is increased in the presence of oxidative stress and also the commonly used anti-fungal drug Flucytosine. Given these results, in my first aim I will determine the rates of switching in C. albicans in response t oxidative stress, nitrosative stress and anti-fungal drug exposure. These factors are common challenges to C. albicans in the clinical setting, and as such, several clinical isolates will be analyzed. I will also identify the response pathway by which they increase phenotypic switching. I hypothesize that the thioredoxin pathway and Rad53 pathways are involved, so they will be tested using gene deletion and over-expression strains for TRX1, TRR1, CAP1, and RAD53. My second aim will address inter-species events between Candida albicans and the fungal and bacterial members of the human oral microbiome. Results from my laboratory shows that pheromone signals from other Candida species induce biofilm formation by C. albicans, which I will further investigate in this aim (14). First, I will determine if signaling between Candida species occurs at physiologically-relevant pheromone concentrations using a set of C. albicans strains I have constructed that secrete pheromones native to these other species in place of its own. I anticipate that my experiments will show that both mating and biofilm formation occur in response to signaling via several Candida species' pheromones, implicating inter-fungal signaling in pathogenesis. Furthermore, I will directly co-culture C. albicans with known bacterial members of the oral microbiome, including Streptococcus gordonii, which secretes high levels of hydrogen peroxide, a known stressor and switch inducer (43). Such results from my second aim will be supported by my first aim. Understanding the white-opaque switch and inter-species signaling events will allow the development of new treatments for preventing the often debilitating, and sometimes even fatal, infections associated with this fungal pathogen.