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

基本信息

批准号：
8777942
负责人：
Gregory M Gauthier
金额：
$ 18.81万
依托单位：
UNIVERSITY OF WISCONSIN-MADISON
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：
2013-12-01 至 2016-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8777942
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 response transcription factor transmission process

项目摘要

DESCRIPTION (provided by applicant): 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.

描述（由申请人提供）：全球，二态真菌每年引起几百万个感染。这些真菌经历了酵母（37oC）和霉菌（22oc）之间的可逆过渡。随着酵母的增长，促进宿主免疫引起疾病的逃避，而随着霉菌的生长促进了土壤中的生存，通过性繁殖通过遗传多样性以及向新宿主传播。尽管热二态性的重要性很重要，但真菌如何调节温度适应的问题知之甚少，并且代表了知识的主要差距。长期目标是描绘真菌适应温度的分子机制。该研究提出，研究了Blastomyces Dermatitidis中的GATA转录因子，SROB（辅助剂量生物合成抑制剂）如何控制温度的适应性，以及该调节是否与铁稳态有关。 SREB无效突变体无法完成22oC的温度依赖性转化率，并且无法正确调节铁稳态。尽管大多数研究都集中在22oC到37oC的温度变化上，但另一个方向的转移-37oC到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）仍未定义。使用基因表达微阵列对SREB进行分析表明，该基因的缺失导致37oC和22oC的转录变化。用定量实时PCR（CHIP-QPCR）染色质免疫沉淀显示SREB在37oC和22oC时结合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”基因进行功能测试。该假设是SREB在GATA基序上结合DNA以调节基因转录，从而控制了对温度的适应，而温度表现为霉菌的过渡。 AIM 1：鉴定基因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。当与基因表达微阵列和基序分析集成时，CHIP-Seq将提供有关SREB如何影响37oC和22oC转录的新的深入知识。 AIM 2：在功能上测试SREB结合的基因，我们具有“手中”（由CHIP-SEQ鉴定的基因）对温度适应的影响（即转换为霉菌）。候选基因“手中”将通过改变转录本丰度并分析从37oC到22oC的过渡期间进行测试。通过CHIP-SEQ确定的其他SREB结合基因（“铁”和“非铁”）将以类似的方式优先考虑和测试。这项研究具有创新性，因为我们专注于二态二态性但不可或缺的一部分，以了解真菌如何适应温度的过渡。这项研究很重要，因为结果将提供新颖的见解，并为真菌用于适应温度的破译机制提供基础。温度适应的基础研究具有长期潜力，可以针对真菌感染患者阐明新的治疗策略。