Molecular Analysis of Mating Type Loci in Heterothallic and Homothallic Cochliobolus spp

异宗和同宗 Cochliobolus spp 交配型基因座的分子分析

• 批准号: 9604856
• 负责人: B. Gillian Turgeon
• 金额: --
• 依托单位: Cornell Univ - State: AWDS MADE PRIOR MAY 2010
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-10-01 至 2001-09-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9604856&HistoricalAwards=false
• 关键词: Molecular; Analysis; Mating; Type; Loci

Turgeon 9604856 Filamentous fungi typically reproduce by both sexual and asexual processes. Sexual ascomycetes may be either heterothallic (require a partner for mating) or homothallic (mate without a partner). The term homothallic as used for filamentous fungi means that a single individual is capable of completing the sexual cycle, usually, but not always, because each nucleus carries both mating type (MAT) genes. In the yeasts, homothallism refers to a change in mating type in some of the cells within a culture of a formerly uniform mating type, followed by mating of "switched" cells with "unswitched" cells. Certain genera of filamentous ascomycetes contain both heterothallic and homothallic species. One of these, Cochliobolus, a pathogen of maize, has been chosen for this study because of its natural diversity of life styles (asexual vs. sexual, heterothallic vs. homothallic, saprophytic vs. parasitic), ready availability of field isolates from a variety of natural habitats and geographical locations, and amenability to molecular genetic manipulation. Like other ascomycetes, the type species (heterothallic C. heterostrophus) has a single mating type locus (MAT) with alternate forms (called idiomorphs and designated MAT-1 and MAT-2) which are each about a kilobase in size, unrelated at the nucleotide level, and encode different transcriptional regulators. The idiomorphs are highly conserved among all Cochliobolus species; in heterothallic species each strain carries a single MAT idiomorph, whereas in homothallics each strain has both idiomorphs within about a kilobase of each other. The sequences on both flanks of the idiomorphs are nearly identical between mating types of the same species and well conserved among species. However, it has recently been found in Neuropora species that one flank of the idiomorph carries a variable region which is about twice as large (7 kb) in homothallic species as in heterothallic (3-5 kb). In addition, the variable region is adjacent to a short ( 57-59bp) constant region which is highly conserved in both mating types and among all species. Although the function of the MAT idiomorphs is clear, the significance of the variable and constant regions is unknown and not easily tested in Neuropora because the genus is not readily amenable to efficient gene targeting. Preliminary evidence suggests variable and constant regions in Cochliobolus analogous to those in Neurospora. One goal of this project is to utilize the efficient gene-targeting ability of Cochilobolus to investigate the functions of these regions: are they important in mating, in the homothallic vs. heterothallic life style, in speciation, or are they dispensable for all of these processes? The analyses will be performed by a combination of sequence analyses, heterologous expression in various genetic backgrounds and by specific gene deletions, replacements, or nucleotide substitutions. Another goal is to construct a phylogeny based on sequences of MAT idiomorphs and flanking regions from a large number of heterothallic and homothallic strains. The intention is to track the evolutionary histories of the various Cochliobolus life styles mentioned above. A practical benefit of the project will be the demonstration, if successful, that a homothallic species can be converted to heterothallic by differential disruption of its MAT genes. This would facilitate genetic analysis of homothallic plant-pathogenic fungal species by forcing crosses and eliminating selfs.

丝状真菌，通常通过有性和无性繁殖。性子囊菌可以是异囊菌（需要伴侣交配）或同囊菌（没有伴侣交配）。“同thallic”一词用于丝状真菌意味着单个个体能够完成性循环，通常，但并非总是如此，因为每个细胞核都携带两种交配型（MAT）基因。在酵母中，同体化是指在以前统一的交配类型的培养物中，一些细胞的交配类型发生变化，随后“切换”的细胞与“未切换”的细胞交配。某些丝状子囊菌属同时含有同种和异囊种。其中一种玉米病原菌——玉米锥虫（Cochliobolus）之所以被选为本研究的病原菌，是因为其生活方式的自然多样性（无性与有性、异源与同源、腐生与寄生）、从各种自然栖息地和地理位置现成的田间分离株，以及易于进行分子遗传操作。像其他子囊菌一样，模式种（异thallic C. heterostrophus）有一个单一的交配型位点（MAT）和其他形式（称为自形，命名为MAT-1和MAT-2），每个都是大约1000个碱基大小，在核苷酸水平上不相关，编码不同的转录调节因子。自形花序在所有种中高度保守；在异thallic种中，每个菌株携带一个单一的MAT自形态，而在同thallic中，每个菌株在彼此约千碱基内具有两个自形态。在同一种的交配类型中，自胚两侧的序列几乎相同，并且在种间具有良好的保守性。然而，最近在Neuropora物种中发现，同thallic物种的一侧翼携带一个可变区域（7 kb），大约是异thallic物种（3-5 kb）的两倍。此外，可变区与一个短的（57-59bp）恒定区相邻，该恒定区在两种交配类型和所有物种中都高度保守。虽然MAT自形的功能是明确的，但可变和恒定区域的意义是未知的，并且不容易在神经孢子虫中进行测试，因为该属不容易适应有效的基因靶向。初步证据表明，耳蜗的可变区和恒定区类似于神经孢子虫。该项目的一个目标是利用Cochilobolus高效的基因靶向能力来研究这些区域的功能：它们在交配、同邻体与异邻体的生活方式、物种形成中是否重要，或者它们在所有这些过程中都是可有可无的？分析将通过序列分析、不同遗传背景的异源表达和特定基因缺失、替换或核苷酸替换的组合来进行。另一个目标是基于大量异thallic和homothallic菌株的MAT自形和侧翼区域序列构建系统发育。目的是追踪上述各种耳蜗生活方式的进化史。这个项目的一个实际好处是，如果成功的话，它将证明同质物种可以通过对其MAT基因的不同破坏而转化为异质物种。这将通过强迫杂交和消除自我来促进同源植物病原真菌物种的遗传分析。