Quantitative genetic approaches to Candida albicans pathogenesis

白色念珠菌发病机制的定量遗传学方法

• 批准号: 10615616
• 负责人: Matthew Z Anderson
• 金额: --
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-12 至 2023-08-01
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10615616
• 关键词: Adherence; Adhesions; Agar; Benign; Biological; Biology; Blood Circulation; Candida; Candida albicans; Candidate Disease Gene; Candidiasis; Cell physiology; Cells; Clinical; Detection; Development; Disease; Disease Outcome; Disparate; Endocytosis Induction; Epithelial Cells; Epithelium; Equilibrium; Esophagus; Filament; Gastrointestinal tract structure; Gene Expression; Genes; Genetic; Genetic Determinism; Genetic Polymorphism; Genetic Screening; Genetic Transcription; Genetic Variation; Growth; HIV; Human; Hyphae; Immune response; Immunocompetent; Immunocompromised Host; In Vitro; Incubated; Individual; Integration Host Factors; Invaded; Investigation; Life; Link; Liquid substance; Maps; Measures; Mucous Membrane; Mus; Nutritional; Oral; Oral candidiasis; Oral cavity; Oral mucous membrane structure; Oropharyngeal; Partner in relationship; Pathogenesis; Pathogenicity; Pathway interactions; Patients; Penetration; Phagocytes; Phenotype; Ploidies; Predisposition; Process; Production; Property; Quality of life; Quantitative Genetics; Quantitative Trait Loci; Recombinants; Role; Signal Transduction; Skin; Solid; Surface; Symbiosis; System; Systemic disease; Systemic infection; Testing; Tissues; Variant; Virulence; Work; Yeasts; cell growth regulation; cell injury; clinically relevant; differential expression; experimental study; fitness; fungus; genetic approach; genome-wide; human pathogen; in vivo; in vivo evaluation; insight; interest; migration; mouse model; mutant; novel; opportunistic pathogen; oral cavity epithelium; oral infection; oropharyngeal thrush; pathogenic fungus; programs; response; therapeutic target; tissue culture; trait; transcriptome sequencing; uptake; urogenital tract; virtual; wasting

ABSTRACT Candida albicans is a common fungal commensal and opportunistic pathogen occupying the skin, oral cavity, gastrointestinal tract, and genitourinary tract of its human host. Overgrowth of C. albicans within these host niches can lead to mucosal and systemic disease and typically relies on pathogenic processes such as the yeast-hyphal transition. Hyphae are capable of breaching host mucosal barriers through active penetration and endocytic uptake that both lead to host cell destruction and transmigration of C. albicans to deeper tissues. Thus, host epithelial cells serve a vital role in detecting C. albicans overgrowth and signaling to professional phagocytes to remove invasive fungal cells. Interestingly, natural C. albicans isolates obtained from clinical setting display a range of filamentation phenotypes, including strains unable to produce hyphae. Here, we will investigate the genetic determinants underlying natural occurring phenotypic variation in C. albicans that regulates the ability to filament and/or colonize the oral cavity through the first use of quantitative genetic approaches in C. albicans. A genetically diverse set of sequenced clinical C. albicans isolates forms the basis of this study as they display significant variation in filamentation processes in vitro and the ability to cause disease in vivo. First, gene expression across the sequenced isolates will facilitate construction of co- expression network modules that associate with in vitro filamentation phenotypes. Expression of key transcriptional regulators within each predicted module will be tested in multiple strain backgrounds for altered in vitro filamentation across a variety of solid and liquid substrates. Automated phenotyping of filamentation, adhesion, and invasion of agar substrates will be built to facilitate scoring mutant phenotypes (Aim 1). In Aim 2, quantitative trait loci (QTL) mapping will be developed utilizing the parasexual program, an alternative mating and ploidy reduction system in C. albicans, to identify the genes responsible for differences in filamentation and epithelial damage between C. albicans strains incubated with OKF6/TERT-2 oral epithelial cells. Identified genes that modulate filamentation and epithelial damage in tissue culture systems will be subsequently tested for in vivo colonization and filamentation phenotypes of a murine model of oropharyngeal candidiasis. Preliminary experiments suggest that filamentation and damage of oral epithelial cells are separable phenotype, which will be explored further through dual-species RNA sequencing of strains separated by filamentation (+/-) and cell damage (+/-) phenotypes incubated with OKF6/TERT-2 cells (Aim 3). This Aim will also determine the role of recently identified host genes with differential expression between damaging and colonizing strains in host cell signaling and transcriptional responses that lead to either filamentation or host cell damage during interaction with oral epithelial cells. Together, these studies will determine the pathways and processes modulated by genetic diversity in C. albicans that lead to divergent filamentation and epithelial damage phenotypes that contribute to commensalism v. disease.

摘要 白色念珠菌是一种常见的真菌性寄生菌和条件致病菌， 腔、胃肠道和泌尿生殖道。C.过度生长白色念珠菌在这些 宿主小生境可导致粘膜和全身性疾病，并且通常依赖于致病过程， 酵母-菌丝过渡。菌丝能够通过主动渗透突破宿主粘膜屏障 和内吞摄取，这两者都导致宿主细胞的破坏和C.白色念珠菌到更深的组织。 因此，宿主上皮细胞在检测C.白色念珠菌过度生长和专业信号 吞噬细胞以去除侵袭性真菌细胞。有趣的是，天然C。从临床获得的白色念珠菌分离株 设置显示了一系列的表达表型，包括不能产生菌丝的菌株。在这里，我们将 研究自然发生的表型变异的遗传决定因素。白色念珠菌， 通过第一次使用定量遗传学方法调节细丝和/或定殖口腔的能力， 方法在C.白色念珠菌。一组遗传多样的测序临床C。白色念珠菌分离物构成了 因为它们在体外表达过程中显示出显著的变化，并且能够引起 体内疾病首先，跨测序分离株的基因表达将有助于构建共- 与体外表达表型相关的表达网络模块。键的表达式 每个预测模块内的转录调节因子将在多个菌株背景中测试，以改变 在各种固体和液体基质上的体外表达。自动化的表型鉴定， 粘附和侵入琼脂基质，以便于对突变体表型进行评分（目标1）。在目标2中， 数量性状基因座（QTL）定位将利用准有性程序，一种替代交配 和C.白念珠菌，以确定基因的差异， C.白色念珠菌菌株与OKF 6/TERT-2口腔上皮细胞孵育。识别 随后将测试在组织培养系统中调节表达和上皮损伤的基因 用于口咽念珠菌病小鼠模型的体内定殖和增殖表型。 初步实验表明，口腔上皮细胞的增殖和损伤是可分离的 表型，这将通过分离菌株的双物种RNA测序进一步探索， 与OKF 6/TERT-2细胞孵育的细胞增殖（+/-）和细胞损伤（+/-）表型（Aim 3）。这一目标将 还确定了最近鉴定的宿主基因的作用，这些基因在损伤和 在宿主细胞信号传导和转录反应中的定殖菌株，其导致表达或宿主 在与口腔上皮细胞相互作用期间的细胞损伤。总之，这些研究将确定 以及C.白色念珠菌，导致分歧的表达和上皮 损害表型，导致过敏症。