Control and Function of Chromatin-Mediated Gene Silencing in Fusarium

镰刀菌染色质介导的基因沉默的控制和功能

• 批准号: 1515998
• 负责人: Michael Freitag
• 金额: $ 57万
• 依托单位: Oregon State University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-08-15 至 2019-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1515998&HistoricalAwards=false
• 关键词: Control; Function; Chromatin; Mediated; Gene

Normal development relies on regulated gene expression, which in many organisms is achieved by balanced activity of protein complexes that turn genes on or off at the right time and in the right place. A major unsolved question is how silencing proteins locate and turn off their target genes. The focus of this research is to discover how genes are targeted and inactivated by the silencing protein complex known as the "polycomb group" (PcG) complex. As a study system, the project will use the fungus, Fusarium graminearum, which has economic importance as a source of beneficial natural products and biofuels and agronomic importance as a plant pathogen. Thus, the results of the research could translate into better methods for manipulating the beneficial traits of the fungus, while minimizing its negative impact on crop plants. The project will have educational impacts by providing research training opportunities for underserved students, laboratory and professional development workshops for high school science teachers, and science presentations to the general public.This project addresses chromatin-based gene silencing by PcG proteins in a novel model organism, the filamentous fungus Fusarium graminearum. The work will take advantage of the fact that Fusarium uses one Polycomb Repressive Complex2 (PRC2) to deliver histone H3 lysine 27 trimethylation (H3K27me3) to specific genes. Preliminary data suggest that the gene recognition is mediated by PRC2 subunits or by interacting proteins and that the subsequently modified genes are sequestered as transcriptionally silent "PRC2 bodies" with clear boundaries to euchromatin. The main objectives of this project are to uncover the mechanism for PRC2 targeting and to define the composition of PRC2 complexes. The aims will be achieved by multi-faceted approaches, including: forward and reverse genetics by undirected selections and screens, gene deletions, and site-directed mutagenesis; biochemistry by chromatin immunoprecipitation, genome-wide expression analyses and mass spectrometry; and cytology by examination of PRC2 bodies in association with DNA. In combination, these techniques will allow identification and characterization of PRC2 signals and proteins involved in gene silencing. Novel components of PRC2-mediated gene silencing in Fusarium will be characterized, and distribution of H3K27me3 and PRC2 in normal and mutant strains will be mapped. The research will provide key knowledge on PRC2-mediated silencing in eukaryotes.

正常发育依赖于基因表达的调节，在许多生物体中，这是通过蛋白质复合物的平衡活性来实现的，这些蛋白质复合物在正确的时间和正确的位置打开或关闭基因。 一个主要的未解决的问题是沉默蛋白如何定位和关闭它们的靶基因。 这项研究的重点是发现基因是如何被沉默蛋白复合物（称为“polycomb group”（PcG）复合物）靶向和灭活的。 作为一个研究系统，该项目将使用真菌，禾谷镰刀菌，它具有经济重要性，作为有益的天然产品和生物燃料的来源，以及作为植物病原体的农学重要性。 因此，研究结果可以转化为更好的方法来操纵真菌的有益性状，同时最大限度地减少其对农作物的负面影响。该项目将通过为服务不足的学生提供研究培训机会，为高中科学教师提供实验室和专业发展研讨会，并向公众提供科学演示，从而产生教育影响。该项目涉及一种新的模式生物，丝状真菌禾谷镰刀菌中基于染色质的基因沉默。这项工作将利用镰刀菌使用一个Polycomb Repressive Complex 2（PRC 2）将组蛋白H3赖氨酸27三甲基化（H3 K27 me 3）传递到特定基因的事实。初步数据表明，基因识别介导的PRC 2亚基或相互作用的蛋白质和随后修饰的基因被隔离的转录沉默的“PRC 2体”与常染色质的明确边界。该项目的主要目标是揭示PRC 2靶向的机制并定义PRC 2复合物的组成。这些目标将通过多方面的方法来实现，包括：正向和反向遗传学，通过非定向选择和筛选，基因缺失和定点诱变;生物化学，通过染色质免疫沉淀，全基因组表达分析和质谱;细胞学，通过检查与DNA相关的PRC 2体。结合这些技术，将允许PRC 2信号和参与基因沉默的蛋白质的鉴定和表征。新的PRC 2介导的基因沉默镰刀菌的组成部分将被表征，并在正常和突变株中的H3 K27 me 3和PRC 2的分布将被映射。这项研究将为真核生物中PRC 2介导的沉默提供关键知识。