Rodríguez-Romero J, Franceschetti M, Bueno E, Sesma A
Centre for Plant Biotechnology and Genomics (CBGP), Universidad Politécnica de Madrid, Campus de Montegancedo, 28223 Pozuelo de Alarcón, Madrid, Spain.
Disease & Stress Biology Department, John Innes Centre, Colney lane, Norwich NR4 7UH, UK.
Nucleic Acids Res. 2015 Jan;43(1):179-95. doi: 10.1093/nar/gku1297. Epub 2014 Dec 16.
Cleavage factor I (CFI) proteins are core components of the polyadenylation machinery that can regulate several steps of mRNA life cycle, including alternative polyadenylation, splicing, export and decay. Here, we describe the regulatory mechanisms that control two fungal CFI protein classes in Magnaporthe oryzae: Rbp35/CfI25 complex and Hrp1. Using mutational, genetic and biochemical studies we demonstrate that cellular concentration of CFI mRNAs is a limited indicator of their protein abundance. Our results suggest that several post-transcriptional mechanisms regulate Rbp35/CfI25 complex and Hrp1 in the rice blast fungus, some of which are also conserved in other ascomycetes. With respect to Rbp35, these include C-terminal processing, RGG-dependent localization and cleavage, C-terminal autoregulatory domain and regulation by an upstream open reading frame of Rbp35-dependent TOR signalling pathway. Our proteomic analyses suggest that Rbp35 regulates the levels of proteins involved in melanin and phenylpropanoids synthesis, among others. The drastic reduction of fungal CFI proteins in carbon-starved cells suggests that the pre-mRNA processing pathway is altered. Our findings uncover broad and multilayer regulatory mechanisms controlling fungal polyadenylation factors, which have profound implications in pre-mRNA maturation. This area of research offers new avenues for fungicide design by targeting fungal-specific proteins that globally affect thousands of mRNAs.
切割因子I（CFI）蛋白是聚腺苷酸化机制的核心组成部分，可调节mRNA生命周期的多个步骤，包括可变聚腺苷酸化、剪接、输出和降解。在这里，我们描述了控制稻瘟病菌中两类真菌CFI蛋白的调控机制：Rbp35/CfI25复合物和Hrp1。通过突变、遗传和生化研究，我们证明CFI mRNA的细胞浓度是其蛋白质丰度的有限指标。我们的结果表明，几种转录后机制在稻瘟病菌中调节Rbp35/CfI25复合物和Hrp1，其中一些在其他子囊菌中也保守。关于Rbp35，这些机制包括C末端加工、RGG依赖的定位和切割、C末端自调节结构域以及Rbp35依赖的TOR信号通路的上游开放阅读框的调节。我们的蛋白质组学分析表明，Rbp35调节参与黑色素和苯丙烷类化合物合成等的蛋白质水平。碳饥饿细胞中真菌CFI蛋白的急剧减少表明前体mRNA加工途径发生了改变。我们的发现揭示了控制真菌聚腺苷酸化因子的广泛和多层调控机制，这对前体mRNA成熟具有深远影响。该研究领域通过靶向全局影响数千个mRNA的真菌特异性蛋白，为杀菌剂设计提供了新途径。
