Two Cdc2 Kinase Genes with Distinct Functions in Vegetative and Infectious Hyphae in Fusarium graminearum.
Two Cdc2 Kinase Genes with Distinct Functions in Vegetative and Infectious Hyphae in Fusarium graminearum.
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DOI：
10.1371/journal.ppat.1004913
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发表时间：
2015-06
期刊：
影响因子：
6.7
通讯作者：
Xu JR
中科院分区：
文献类型：
--
作者：
Liu H;Zhang S;Ma J;Dai Y;Li C;Lyu X;Wang C;Xu JR
真核细胞周期涉及许多在有丝分裂过程中起重要作用的蛋白激酶，其中大多数在芽殖酵母和分裂酵母中都有很好的特征，在其他真菌中也是保守的。然而，与模式酵母和丝状真菌不同，模型酵母和丝状真菌只有一个细胞周期所必需的CDC2，而小麦赤霉菌Fusum graminearum包含两个CDC2同源基因。突变体cdc2A和cdc2B在生长速度和分生孢子形成方面没有明显的缺陷，但两者的缺失都是致命的，这表明这两个CDC2同源基因在营养生长和无性繁殖过程中具有多余的功能。然而，虽然cdc2B突变体是正常的，但cdc2A突变体的毒力显著降低，很少产生子囊孢子。虽然cdc2a缺失对小麦侵染枝条或下胚轴的形成没有明显影响，但cdc2a突变体在小麦组织中侵染生长的分化和生长方面受到限制。因此，CDC2A在感染性生长和有性生殖的细胞周期调控中发挥着阶段特异性的作用。在植物侵染和子囊孢子发生过程中，CDC2A和CDC2B都有结构性表达，但只有CDC2A表达上调。在营养菌丝、子囊孢子和侵染菌丝中观察到CDC2A-GFP定位于细胞核，而不定位于CDC2B-GFP。嵌合融合构建体的互补分析表明，CDC2AN-末端和C-末端在致病和子囊孢子发生中都是重要的，但只有N-末端对于其亚细胞定位是重要的。在菌丝酵母菌中，与禾谷镰刀菌关系密切的只有3种镰刀菌有两个CDC2基因。此外，禾谷镰刀菌独有的两个Aurora激酶基因和一个额外的可能的细胞周期蛋白基因，其在萌芽酵母中与CAK1和其他四种必需的有丝分裂酶的同源基因是生存所必需的。总之，我们的数据表明禾谷镰刀菌营养菌丝和侵染菌丝对细胞周期的调控是不同的，在子囊孢子发生和植物侵染过程中，CDc2A可能通过与一个特定阶段的细胞周期蛋白相互作用发挥着比CDc2B更重要的作用。在模型酵母和丝状真菌中，CDC2是一个必不可少的基因，它编码有丝分裂细胞周期进程中唯一必需的CDK。然而，小麦赤霉菌F.graminearum含有两个CDC2同源基因。Cdc2A和cdc2B缺失突变体在营养生长方面没有缺陷，但两者的缺失都是致命的。而cdc2B突变体是正常的，而cdc2A突变体几乎是非致病的，这表明在侵染菌丝中只有cdc2A是必需的。在有丝分裂活跃的细胞中，CDc2A和CDc2B在亚细胞定位上不同，只有在细胞核中的CDc2A定位增加。此外，禾谷镰刀菌具有两个Ipl1极光激酶基因的同源基因，删除了5个必需的酵母有丝分裂酶基因的同源基因的突变体是可行的。然而，这些突变体中的大多数在毒力上都显著降低。总体而言，我们的数据表明，禾谷镰刀菌在CDK和其他关键有丝分裂酶基因上与模式真菌不同，营养菌丝和侵染菌丝对细胞周期的调控也不同。这是首次报道真菌中的两种CDc2激酶，它们在有性繁殖和植物侵染过程中的亚细胞定位和功能不同。
Eukaryotic cell cycle involves a number of protein kinases important for the onset and progression through mitosis, most of which are well characterized in the budding and fission yeasts and conserved in other fungi. However, unlike the model yeast and filamentous fungi that have a single Cdc2 essential for cell cycle progression, the wheat scab fungus Fusarium graminearum contains two CDC2 orthologs. The cdc2A and cdc2B mutants had no obvious defects in growth rate and conidiation but deletion of both of them is lethal, indicating that these two CDC2 orthologs have redundant functions during vegetative growth and asexual reproduction. However, whereas the cdc2B mutant was normal, the cdc2A mutant was significantly reduced in virulence and rarely produced ascospores. Although deletion of CDC2A had no obvious effect on the formation of penetration branches or hyphopodia, the cdc2A mutant was limited in the differentiation and growth of infectious growth in wheat tissues. Therefore, CDC2A plays stage-specific roles in cell cycle regulation during infectious growth and sexual reproduction. Both CDC2A and CDC2B are constitutively expressed but only CDC2A was up-regulated during plant infection and ascosporogenesis. Localization of Cdc2A- GFP to the nucleus but not Cdc2B-GFP was observed in vegetative hyphae, ascospores, and infectious hyphae. Complementation assays with chimeric fusion constructs showed that both the N- and C-terminal regions of Cdc2A are important for its functions in pathogenesis and ascosporogenesis but only the N-terminal region is important for its subcellular localization. Among the Sordariomycetes, only three Fusarium species closely related to F. graminearum have two CDC2 genes. Furthermore, F. graminearum uniquely has two Aurora kinase genes and one additional putative cyclin gene, and its orthologs of CAK1 and other four essential mitotic kinases in the budding yeast are dispensable for viability. Overall, our data indicate that cell cycle regulation is different between vegetative and infectious hyphae in F. graminearum and Cdc2A, possibly by interacting with a stage-specific cyclin, plays a more important role than Cdc2B during ascosporogenesis and plant infection. In the model yeasts and filamentous fungi, CDC2 is an essential gene that encodes the only CDK essential for mitotic cell cycle progression. However, the wheat scab fungus F. graminearum contains two CDC2 orthologs. The cdc2A and cdc2B deletion mutants had no defects in vegetative growth but deletion of both is lethal. Whereas the cdc2B mutant was normal, the cdc2A mutant was almost non-pathogenic, indicating that only Cdc2A is essential in infectious hyphae. Cdc2A and Cdc2B differ in subcellular localization and only localization of Cdc2A to the nucleus was increased in cells active in mitosis. Furthermore, F. graminearum uniquely has two orthologs of Ipl1 Aurora kinase and mutants deleted of orthologs of five essential yeast mitotic kinase genes were viable. However, most of these mutants were significantly reduced in virulence. Overall, our data indicate that F. graminearum differs from the model fungi in CDK and other key mitotic kinase genes, and cell cycle regulation is different between vegetative and infectious hyphae. This is the first report on two Cdc2 kinases in fungi and they differ in subcellular localization and functions during sexual reproduction and plant infection.
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