Department of Plant Pathology and Microbiology, Robert H. Smith Faculty of Agriculture, Food and Environment, The Hebrew University, Rehovot, 76100, Israel.
Department of Medical Biochemistry and Biophysics, Umeå University, Umeå, Sweden.
DNA Repair (Amst). 2019 Nov;83:102674. doi: 10.1016/j.dnarep.2019.102674. Epub 2019 Jul 24.
Ribonucleotide reductase (RNR) catalyzes the rate limiting step in dNTP biosynthesis and is tightly regulated at the transcription and activity levels. One of the best characterized responses of yeast to DNA damage is up-regulation of RNR transcription and activity and consequently, elevation of the dNTP pools. Hydroxyurea is a universal inhibitor of RNR that causes S phase arrest. It is used in the clinic to treat certain types of cancers. Here we studied the response of the fungal plant pathogen Fusarium oxysporum to hydroxyurea in order to generate hypotheses that can be used in the future in development of a new class of pesticides. F. oxysporum causes severe damage to more than 100 agricultural crops and specifically threatens banana cultivation world-wide. Although the recovery of F. oxysporum from transient hydroxyurea exposure was similar to the one of Saccharomyces cerevisiae, colony formation was strongly inhibited in F. oxysporum in comparison with S. cerevisiae. As expected, genomic and phosphoproteomic analyses of F. oxysporum conidia (spores) exposed to hydroxyurea showed hallmarks of DNA replication perturbation and activation of recombination. Unexpectedly and strikingly, RNR was not induced by either hydroxyurea or the DNA-damaging agent methyl methanesulfonate as determined at the RNA and protein levels. Consequently, dNTP concentrations were significantly reduced, even in response to a low dose of hydroxyurea. Methyl methanesulfonate treatment did not induce dNTP pools in F. oxysporum, in contrast to the response of RNR and dNTP pools to DNA damage and hydroxyurea in several tested organisms. Our results are important because the lack of a feedback mechanism to increase RNR expression in F. oxysporum is expected to sensitize the pathogen to a fungal-specific ribonucleotide inhibitor. The potential impact of our observations on F. oxysporum genome stability and genome evolution is discussed.
核糖核苷酸还原酶 (RNR) 催化 dNTP 生物合成的限速步骤，其转录和活性水平受到严格调控。酵母对 DNA 损伤的最佳特征反应之一是 RNR 转录和活性的上调，以及 dNTP 池的升高。羟基脲是 RNR 的通用抑制剂，可导致 S 期停滞。它在临床上用于治疗某些类型的癌症。在这里，我们研究了真菌植物病原体尖孢镰刀菌对羟基脲的反应，以便提出假设，这些假设可用于未来开发一类新的杀虫剂。尖孢镰刀菌对 100 多种农作物造成严重损害，特别是对全球香蕉种植构成威胁。尽管尖孢镰刀菌从短暂的羟基脲暴露中的恢复与酿酒酵母相似，但与酿酒酵母相比，尖孢镰刀菌的菌落形成受到强烈抑制。正如预期的那样，对羟基脲处理的尖孢镰刀菌分生孢子（孢子）的基因组和磷酸蛋白质组分析显示了 DNA 复制扰动和重组激活的特征。出乎意料的是，在 RNA 和蛋白质水平上，羟基脲或 DNA 损伤试剂甲磺酸甲酯都没有诱导 RNR 的表达。因此，即使在低剂量羟基脲的作用下，dNTP 浓度也显著降低。与几种测试生物中 RNR 和 dNTP 对 DNA 损伤和羟基脲的反应相反，甲磺酸甲酯处理并没有诱导尖孢镰刀菌的 dNTP 池。我们的研究结果很重要，因为在尖孢镰刀菌中缺乏增加 RNR 表达的反馈机制，预计会使病原体对真菌特异性核糖核苷酸抑制剂敏感。我们的观察结果对尖孢镰刀菌基因组稳定性和基因组进化的潜在影响进行了讨论。
