镰刀菌,甘蔗的关键病原体,负责中国的Pokkahboeng病(PBD)。14-3-3蛋白与关键的发育过程有关,包括二态转变,信号转导,和各种植物病原真菌的碳代谢。然而,他们的角色在F.sacchari中知之甚少。这项研究集中于两个14-3-3蛋白编码基因的表征,FsBmh1和FsBmh2,在糖精内。发现这两个基因都在营养生长阶段表达,然而FsBmh1在体外孢子形成阶段被抑制。为了阐明这些基因的功能,产生缺失突变体ΔFsBmh1和ΔFsBmh2。ΔFsBmh2表现出更明显的表型缺陷,如菌丝分支受损,隔膜,分生孢子,孢子萌发,和菌落生长,与ΔFsBMH1相比。值得注意的是,两种敲除突变体均显示出毒力降低,转录组分析揭示了与观察到的表型相关的变化。为了进一步研究FsBmh1和FsBmh2之间的功能相互作用,我们构建并分析了具有组合缺失和沉默(ΔFsBmh/siFsBmh)以及过表达(O-FsBmh)的突变体。ΔFsBmh1/siFsBmh2或ΔFsBmh2/siFsBmh1的组合比单等位基因缺失的组合表现出更严重的表型,表明两个14-3-3蛋白之间的功能冗余。酵母双杂交(Y2H)测定鉴定出20种在初级代谢或多种生物学功能中起关键作用的蛋白质,其中12个与FsBmh1和FsBmh2相互作用。三种蛋白质与FsBmh1特异性相关,而五种仅与FsBmh2相互作用。总之,这项研究为FsBmh1和FsBmh2在糖精中的作用提供了新的见解,并通过调节FsBmh功能突出了PBD管理的潜在目标。
Fusarium sacchari, a key pathogen of sugarcane, is responsible for the Pokkah boeng disease (PBD) in
China. The 14-3-3 proteins have been implicated in critical developmental processes, including dimorphic transition, signal transduction, and carbon metabolism in various phytopathogenic fungi. However, their roles are poorly understood in F. sacchari. This study focused on the characterization of two 14-3-3 protein-encoding genes, FsBmh1 and FsBmh2, within F. sacchari. Both genes were found to be expressed during the vegetative growth stage, yet FsBmh1 was repressed at the
sporulation stage in vitro. To elucidate the functions of these genes, the deletion mutants ΔFsBmh1 and ΔFsBmh2 were generated. The ΔFsBmh2 exhibited more pronounced phenotypic defects, such as impaired hyphal branching, septation, conidiation, spore germination, and colony growth, compared to the ΔFsBmh1. Notably, both knockout mutants showed a reduction in virulence, with transcriptome analysis revealing changes associated with the observed phenotypes. To further investigate the functional interplay between FsBmh1 and FsBmh2, we constructed and analyzed mutants with combined deletion and silencing (ΔFsBmh/siFsBmh) as well as overexpression (O-FsBmh). The combinations of ΔFsBmh1/siFsBmh2 or ΔFsBmh2/siFsBmh1 displayed more severe phenotypes than those with single allele deletions, suggesting a functional redundancy between the two 14-3-3 proteins. Yeast two-hybrid (Y2H) assays identified 20 proteins with pivotal roles in primary metabolism or diverse biological functions, 12 of which interacted with both FsBmh1 and FsBmh2. Three proteins were specifically associated with FsBmh1, while five interacted exclusively with FsBmh2. In summary, this research provides novel insights into the roles of FsBmh1 and FsBmh2 in F. sacchari and highlights potential targets for PBD management through the modulation of FsBmh functions.