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：在世界范围内，二相性真菌每年造成数百万人感染。这些真菌经历了一个 酵母(37摄氏度)和霉菌(22摄氏度)之间的可逆转变。酵母生长促进逃避寄主 免疫导致疾病，而霉菌的生长促进了土壤中的生存，遗传多样性通过有性繁殖 繁殖，并传播给新的宿主。尽管热二形现象很重要，但问题是 真菌是如何调节温度适应的，人们对此知之甚少，这是知识上的一大空白。这个 长期目标是阐明真菌适应温度的分子机制(S)。这项研究 Proposed研究了皮炎芽孢杆菌中的GATA转录因子SREB(铁载体)是如何 生物合成抑制子)，调控对温度的适应，以及这一调节是否 与铁稳态有关。SREB零突变体不能完成依赖温度的转化 霉菌在22摄氏度，不能适当地调节铁的动态平衡。虽然大多数研究都集中在 温度从22摄氏度到37摄氏度的变化，另一个方向的变化-37摄氏度到22摄氏度-被低估了。 此外，下游靶基因和机制用于响应温度(37摄氏度或22摄氏度) 保持模糊的定义。用基因表达芯片分析SREB发现该基因缺失 在37℃和22℃时引起转录的多效性变化。定量染色质免疫沉淀 实时定量聚合酶链式反应(ChIP-qPCR)显示，SREB在37oC和22oC时结合了功能不同的基因。 此外，在SREB控制下的几个候选“非铁”和“铁”基因已经被确定为 功能测试。假设SREB结合GATA基序上的DNA来调节基因转录，这在 反过来，控制对温度的适应，这是通过向模具的过渡所表现出来的。目标1：识别基因 SREB利用带DNA测序的芯片(CHIP-SEQ)在37℃和22℃的全基因组范围内与SREB结合。 CHIP-SEQ是一种高效的方法，可以在不偏向特定基序的情况下识别SREB结合的基因。 当与基因表达微阵列和基序分析相结合时，CHIP-SEQ将提供新的、深入的 关于SREB如何影响37摄氏度和22摄氏度转录的知识。目标2：功能测试SREB绑定 我们手中的基因(以及由CHIP-SEQ识别的基因)对温度适应的影响(即， 转换为模具)。候选基因将通过改变转录丰度进行测试和分析 用于从37摄氏度到22摄氏度的过渡过程中的缺陷。额外的SREB结合基因(“铁”和“非铁”) 将以类似的方式对CHIP-SEQ识别的数据进行优先排序和测试。这项研究具有创新性，因为我们 都集中在一个未被研究的，但不可分割的二形性，过渡到模具了解如何 真菌能适应温度。这项研究具有重要意义，因为其结果将提供新的见解和服务 作为破译真菌适应温度的机制的基础。关于温度的基础研究 适应具有长期的潜力，为真菌感染患者提供新的治疗策略。