Virulence factor identification by comparative transcriptomics in Candida species

通过比较转录组学鉴定念珠菌属毒力因子

基本信息

批准号：
8646883
负责人：
Michael C Lorenz
金额：
$ 19万
依托单位：
UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-04-12 至 2016-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8646883
关键词：
Accounting Animal Model Antifungal Agents Antifungal Therapy Bioinformatics Bone Marrow Candida Candida albicans Candida glabrata Cells Clinical Coculture Techniques Codon Nucleotides Complex Control Animal Cues Data Dendritic Cells Diagnosis Disease Disseminated candidiasis Endothelial Cells Epidemiology Epithelial Cells Event Explosion Frequencies Gene Expression Profiling Genes Genetic Genetic Variation Genome Genomics Glyoxylates Goals Health High-Throughput RNA Sequencing Immune response Immunity Incidence Infection Interferons Knock-out Libraries Life Literature Mammalian Cell Maps Mediating Metabolic Microbe Molecular Mus Mycoses Outcome Pathogenesis Pathway interactions Pattern Phagocytes Phagocytosis RNA RNA Interference RNA Sequences Reading Refractory Relative (related person)Reporting Resistance Resources Rest Role Saccharomyces cerevisiae Side Stress Technology Testing Text Transcript Translating Vaginitis Variant Virulence Virulence Factors Virulent Work base cell type clinically significant comparative fungus genome annotation genome sequencing glyoxylate immunoregulation killings knock-down macrophage mortality mutant neutrophil novel oropharyngeal thrush pathogen public health relevance response tool trait transcriptomics

项目摘要

DESCRIPTION (provided by applicant): Collectively, the genus Candida is the most important cause of fungal infections in the developed world, responsible for roughly 75% of disseminated or invasive fungal infections. This poses a significant clinical challenge as disseminated candidiasis is difficult to diagnose and is often refractory to antifungal therapy, leading to a mortality rate that has remained stubbornly high, at around 40%, for decades. Overlapping the Candida genus is a group of species referred to as the CTG clade, because they translate the CUG codon non-canonically. The CTG clade encompasses all of the clinically significant Candida species with the exception of C. glabrata, a commonly isolated pathogen despite being much more closely related to Saccharomyces cerevisiae than to CTG species, and C. krusei, a rare pathogen. Within the CTG clade there is a wide variation in the frequency with which the species are isolated: precise numbers differ between studies, but C. albicans remains responsible for more than half of disseminated candidiasis infections, while C. tropicalis and C. parapsilosis (~10% each) are commonly isolated. C. lusitaniae and C. guillermondii are infrequent pathogens and C. famata (aka Debaromyces hansenii) is rarely seen. Data from carefully controlled animal models broadly confirm that this epidemiological pattern reflects the inherent virulence of these species, though C. parapsilosis is probably less virulent than its incidence rate would imply. C. albicans is even more dominant in other common manifestations of candiosis, such as vaginitis and oropharyngeal thrush. The genetic diversity encompassed by these related species offers a tool with which to understand virulence in a novel way: by dissecting the response of each of these species to interactions with host cell through transcriptomics. This has been a very successful approach with C. albicans, identifying pathways of both stress resistance and metabolic adaptations. It has also highlighted large numbers of uncharacterized genes, many of which are specific to the CTG clade and some specific only to C. albicans and its closest relatives. We propose here to use comparative transcriptional profiling of seven CTG clade species, with a range of virulence from high (C. albicans) to very low (D. hansenii), during co-cultures with macrophages, a key component of mammalian antifungal immunity. Using high-throughput RNA sequencing (RNA-seq), we will quantitatively determine transcript abundance from both the fungal and mammalian component during these interactions. Bioinformatic analysis will identify genes that are highly induced in th most virulent species (C. albicans and C. tropicalis) but either do not exist or are not regulated in the less virulent species. A prioritized subset of these genes will then be analyzed using molecular approaches to understand their role in virulence. Simultaneously, the profiles of the murine macrophages will both fill a present hole in the literature (no transcriptional analyses of murine primary cells have been reported) and, more importantly, will identify how the most virulent species may blunt the typical antifungal response to promote survival; several mechanisms of immunomodulation have been proposed but very little is known about how they might work. Finally, the clinical prominence of C. albicans has led to the vast majority of molecular work being performed in this single species. Far more genes have been knocked out in C. albicans than in all the other CTG species combined, even before considering mutant libraries generated by several labs. Very few transcript profiling studies have been performed in non-albicans species, and the data generated by this project will be an extremely valuable resource to understand these important species and to contribute to annotation of these genomes.

描述（由申请人提供）：统称，念珠菌属是发达国家真菌感染的最重要原因，约有75％的传播或侵入性真菌感染。这构成了重大的临床挑战，因为很难诊断出散布的念珠菌病，并且通常对抗真菌治疗难治性，导致死亡率一直保持高，数十年来约40％。念珠菌属重叠是一组称为CTG进化枝的物种，因为它们非统一地翻译了CUG密码子。 CTG进化枝涵盖了所有具有临床意义的念珠菌物种，除了glabrata（尽管与酿酒酵母的相关性密切），但通常是一种孤立的病原体，而与CTG物种的相关性更大。在CTG进化枝内，该物种分离的频率存在很大差异：研究之间的精确数量有所不同，但是白色念珠菌仍负责超过一半的散布念珠菌感染，而Tropicalis和C. parapappicalis和C. parapapapaplisois（每个）通常是分离的。 C. lusitaniae和C. guillermondii是很少发生的病原体，而C. famata（又名Hansenii）很少见。经过精心控制的动物模型的数据广泛证实，这种流行病学模式反映了这些物种的固有毒力，尽管副细胞链球菌可能比其发病率所暗示的毒素不那么毒。白色念珠菌在其他常见的念珠菌（例如阴道炎和口咽鹅口疮）中更为主导。这些相关物种包含的遗传多样性提供了一种以新颖方式理解毒力的工具：通过解剖这些物种对通过转录组学与宿主细胞相互作用的反应。对于白色念珠菌，这是一种非常成功的方法，可以确定抗压力和代谢适应的途径。它还强调了大量未表征的基因，其中许多特定于CTG进化枝，有些仅针对白色念珠菌及其最接近的亲属。我们在这里提议在与巨噬细胞的共培养期间，使用七个CTG进化枝种的比较转录分析，从高（白色念珠菌）到非常低的毒力，这是巨噬细胞的共培养，这是哺乳动物抗真菌免疫的关键组成部分。使用高通量RNA测序（RNA-Seq），我们将在这些相互作用期间定量确定来自真菌和哺乳动物成分的转录本丰度。生物信息学分析将鉴定出在大多数毒性物种（白色念珠菌和热带梭菌）中高度诱导的基因，但在不太毒性物种中不存在或不受调节。然后，将使用分子方法来分析这些基因的优先级子集，以了解其在毒力中的作用。同时，鼠巨噬细胞的特征都将填充文献中的当前孔（尚未报告鼠原代细胞的转录分析），更重要的是，将确定最具毒性的物种如何钝化典型的抗真菌反应以促进生存；已经提出了几种免疫调节的机制，但对它们的工作方式知之甚少。最后，白色念珠菌的临床突出导致在该物种中进行的绝大多数分子作品。甚至在考虑了几个实验室产生的突变库之前，在白色念珠菌中，基因被淘汰的基因被淘汰了。在非阿尔比亚物种中，很少有成绩单分析研究进行，该项目产生的数据将是一种非常宝贵的资源，可以理解这些重要物种并为这些基因组的注释做出贡献。