Regulation of Morphogenesis in Dimorphic Fungi by a GATA Transcription Factor

GATA 转录因子对二态真菌形态发生的调节

• 批准号: 8636380
• 负责人: Gregory M Gauthier
• 金额: $ 22.58万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-12-01 至 2015-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8636380
• 关键词: Affect; Anabolism; Animal Model; Basic Science; Binding; Binding Sites; Biology; Blastomyces; Blastomyces dermatitidis; Candidate Disease Gene; Categories; Cryptococcus neoformans; DNA; DNA Binding; DNA Sequence; DNA Sequence Analysis; Data; Defect; Development; Disease; Event; Foundations; Gene Chips; Gene Deletion; Gene Expression Microarray Analysis; Gene Targeting; Generations; Genes; Genetic Transcription; Genetic Variation; Goals; Growth; Hand; Health; Histoplasma capsulatum; Homeostasis; Homologous Gene; Hour; Human; Immune system; Immunity; Infection; Iron; Knowledge; Life Style; Link; Molds; Molecular; Morphogenesis; Morphology; Mycoses; Ontology; Oxidation-Reduction; Partner in relationship; Pathogenesis; Patients; Persons; Phase Transition; Pneumonia; Process; Regulation; Reporter; Reproduction; Research; Saccharomycetales; Siderophores; Soil; Staging; Temperature; Testing; Therapeutic; Time; Transcript; Virulent; Yeasts; chromatin immunoprecipitation; dimorphism; fatty acid metabolism; fungus; genome-wide; human tissue; in vivo; innovation; insight; mutant; novel; novel therapeutics; overexpression; pathogen; public health relevance; response; transcription factor; transmission process

ABSTRACT: Worldwide, the dimorphic fungi cause several million infections each year. These fungi undergo a reversible transition between yeast (37oC) and mold (22oC). Growth as yeast promotes evasion of host immunity to cause disease, whereas growth as mold promotes survival in soil, genetic diversity through sexual reproduction, and transmission to new hosts. Despite the importance of thermal dimorphism, the question of how fungi regulate temperature adaptation is poorly understood and represents a major gap in knowledge. The long-term goal is to delineate the molecular mechanism(s) used by fungi to adapt to temperature. The research proposed investigates how a GATA transcription factor in Blastomyces dermatitidis, SREB (siderophore biosynthesis repressor in Blastomyces), governs the adaptation to temperature, and whether this regulation is linked with iron homeostasis. SREB null mutants fail to complete the temperature-dependent conversion to mold at 22oC and cannot properly regulate iron homeostasis. While most research has focused on the temperature change from 22oC to 37oC, the shift in the other direction - 37oC to 22oC - is underappreciated. Moreover, the downstream target genes and mechanisms used to respond to temperature (37oC or 22oC) remain ill defined. Analysis of SREB using gene expression microarrays revealed that deletion of this gene caused pleiotropic changes in transcription at 37oC and 22oC. Chromatin immunoprecipitation with quantitative real-time PCR (ChIP-qPCR) demonstrated SREB binds genes with disparate functions at 37oC and 22oC. Moreover, several candidate "non-iron" and "iron" genes under the control of SREB have been identified for functional testing. The hypothesis is SREB binds DNA at GATA motifs to regulate gene transcription, which in turn, controls the adaptation to temperature that is manifested by the transition to mold. Aim 1: Identify genes SREB binds in vivo on a genome-wide scale at 37oC and 22oC using ChIP with DNA sequencing (ChIP-seq). ChIP-seq is highly efficient and allows identification of SREB-bound genes without bias to specific motifs. When integrated with gene expression microarray and motif analyses, ChIP-seq will provide new, in-depth knowledge about how SREB impacts transcription at 37oC and 22oC. Aim 2: Functionally test SREB-bound genes we have "in-hand" (and those identified by ChIP-seq) for their impact on temperature adaptation (i.e., conversion to mold). Candidate genes "in-hand" will be tested by altering transcript abundance and analyzed for defects during the transition from 37oC to 22oC. Additional SREB-bound genes ("iron" and "non-iron") identified by ChIP-seq will be prioritized and tested in a similar fashion. The research is innovative because we are focusing on an understudied, but integral part of dimorphism, the transition to mold to understand how fungi adapt to temperature. The research is significant because the results will provide novel insight and serve as a foundation to decipher mechanisms used by fungi to adapt to temperature. Basic research on temperature adaptation has long-term potential to illuminate new therapeutic strategies for patients with fungal infections.

摘要：全球，二态真菌每年引起数百万个感染。这些真菌经历了 酵母（37oC）和霉菌（22oC）之间的可逆过渡。随着酵母促进寄主的生长 免疫可引起疾病，而随着霉菌的生长促进土壤中的生存，遗传多样性通过性促进 繁殖，并传输到新宿主。尽管热二态性很重要，但 真菌如何调节温度适应的理解很少，并且代表了知识的主要差距。这 长期目标是描述真菌用于适应温度的分子机制。研究 拟议的研究调查了Blastomyces Dermatitidis中的GATA转录因子如何SREB（铁载体 爆炸性的生物合成抑制剂），控制温度的适应，以及该调节是否为 与铁稳态有关。 SREB无效突变体无法完成与温度有关的转化 22oC的霉菌，无法正确调节铁稳态。虽然大多数研究都集中在 温度从22oC到37oC，另一个方向的转移-37oC向22oC-被低估了。 此外，用于响应温度的下游靶基因和机制（37oC或22oC） 保持不明的定义。使用基因表达微阵列对SREB进行分析表明，该基因的缺失 在37oC和22oC处引起转录的多效变化。定量染色质免疫沉淀 实时PCR（CHIP-QPCR）表现出SREB在37oC和22oC处具有不同功能的基因。 此外，已经确定了在SREB控制下的几个候选“非铁”和“铁”基因 功能测试。该假设是SREB在GATA基序上结合DNA以调节基因转录，该基因在 转弯，控制对温度的适应，这表现为霉菌的过渡。目标1：识别基因 SREB使用与DNA测序（CHIP-SEQ）的芯片在37oC和22oC上在基因组范围内结合体内。 ChIP-Seq高效，可以鉴定出与特定基序没有偏置的SREB结合基因。 与基因表达微阵列和基序分析集成时，Chip-Seq将提供新的，深入的 有关SREB如何影响37oC和22oC的转录的知识。 AIM 2：功能测试SREB结合 我们的基因具有“手中”（以及chip-seq确定的基因）对温度适应的影响（即 转换为霉菌）。候选基因“手中”将通过改变转录物丰度并进行分析来测试 对于从37oC到22oC的过渡期间的缺陷。其他SREB结合基因（“铁”和“非铁”） 通过CHIP-SEQ确定的将以类似的方式进行优先级和测试。这项研究具有创新性，因为我们 专注于二态性的经过研究的但不可或缺的一部分 真菌适应温度。这项研究很重要，因为结果将提供新颖的见解并服务 作为真菌用于适应温度的破译机制的基础。温度基础研究 适应具有长期的潜力，可以针对真菌感染的患者阐明新的治疗策略。