Regulation of Sexual Biofilm Development in Candida albicans

真菌白色念珠菌是人类微生物群的一种共生物种。它也是一种经常分离的机会性病原体，负责各种粘膜和危及生命的全身感染。这些感染中的大多数是由于白色念珠菌细胞存在于生物膜中的能力——具有独特特性的具有弹性的、表面相关的细胞群落。许多严重感染与植入医疗器械高度相关，植入医疗器械为生物膜的形成提供了有效的底物。近年来的研究表明，信息素信号是一种促进白色念珠菌生物膜形成的新机制。信息素信号已被确定为在白色念珠菌细胞之间编排交配。然而，令人惊讶的是，这个程序也可以诱导反应细胞成为粘附并进行生物膜发育。信息素可以在不同交配类型的细胞共培养中驱动生物膜的形成。此外，该途径的自激活甚至可以发生在单性别培养中，这同样会增强白色念珠菌细胞的粘附性和生物膜的发育。因此，信息素可以在多种白色念珠菌细胞类型驱动生物膜形成的过程中发挥多向性作用，包括通过常规和非常规信息素信号通路。在这个提议中，我们试图研究信息素诱导的生物膜形成的机制，并利用两种生物膜感染的哺乳动物模型来解决它们在体内形成的可能性。该提案的两个目标是：明确信息素诱导生物膜形成的机制。我们将确定信息素信号诱导白色念珠菌生物膜形成的机制。在信息素诱导的生物膜形成过程中，将对多种临床分离株进行转录谱分析，并通过CRISPR删除与生物膜形成相关的靶基因，并评估其在这一过程中的作用。目标2。确定信息素信号在体内生物膜形成中的作用。我们将使用两种不同的哺乳动物生物膜形成模型（大鼠导管模型和假牙口腔炎模型）来确定信息素信号通路是否会导致生物膜形成的增强。总之，这些实验的完成将揭示信息素信号通路在体外和体内如何影响白色念珠菌的生物膜形成。我们的方法是基于有趣的初步数据，这些数据表明，在动物模型中引入两种交配类型的混合物会导致生物膜的增强。如果成功，这一建议有可能改变我们对白色念珠菌生物膜形成的理解，并对真菌感染患者的治疗具有直接的临床影响。总的来说，最先进的基因组方法、分子遗传学、高分辨率显微镜、微流体模型和两种经过验证的模拟人类生物膜感染的动物模型的结合，为研究这一重要的临床问题提供了一种强大的组合方法。

The fungus Candida albicans is a commensal species of the human microbiota. It is also a frequently isolated opportunistic pathogen, responsible for a variety of mucosal and life-threatening systemic infections. The majority of these infections are a result of the ability of C. albicans cells to exist in biofilms – resilient, surface-associated, communities of cells with unique properties. Many serious infections are highly correlated with implanted medical devices, which provide efficient substrates for biofilm formation. Recent studies have revealed that pheromone signaling is a novel mechanism promoting the formation of C. albicans biofilms. Pheromone signaling is well established to choreograph mating between C. albicans cells. Surprisingly, however, this program can also induce responding cells to become adherent and undergo biofilm development. Pheromones can drive biofilm formation in co-cultures of cells of opposite mating types. In addition, auto-activation of this pathway can even occur in single sex cultures, which similarly results in enhanced adherence of C. albicans cells and biofilm development. It is therefore clear that pheromones can play a pleiotropic role in driving biofilm formation by multiple C. albicans cell types, both by conventional and non-conventional pheromone signaling pathways. In this proposal, we seek to investigate the mechanism of pheromone-induced biofilm formation, and to address the potential for them to form in vivo using two mammalian models of biofilm infection. The two Aims of the proposal are: Aim 1. To define the mechanism of pheromone-induced biofilm formation. We will determine the mechanism by which pheromone signaling induces biofilm formation in C. albicans. Transcriptional profiling will be performed on a diverse set of clinical isolates during pheromone-induced biofilm formation, and target genes implicated in biofilm formation will be deleted by CRISPR and evaluated for their roles in this process. Aim 2. To determine the role of pheromone signaling on biofilm formation in vivo. We will determine if pheromone-signaling pathways lead to enhanced biofilm formation using two distinct mammalian models of biofilm formation, a rat catheter model and a denture stomatitis model. Together, completion of these experiments will reveal how pheromone-signaling pathways impact biofilm formation in C. albicans both in vitro and in vivo. Our approach is based on intriguing preliminary data that indicates that introduction of a mixture of two mating types results in enhanced biofilms in an animal model. If successful, this proposal has the potential to transform our understanding of C. albicans biofilm formation, and has direct clinical ramifications for treatment of fungal infections in patients. Overall, the combination of state- of-the-art genomic approaches, molecular genetics, high-resolution microscopy, microfluidic models, and two validated animal models designed to mimic human biofilm infections, provides a powerful combinatorial approach to studying this important clinical problem.