PDF(Pubmed)
Abstract：
The p21-GTPase-activated protein kinases (PAKs) participate in signal transduction downstream of Rho GTPases, which are regulated by Rho GTPase-activating proteins (Rho-GAP). Herein, we characterized two orthologous Rho-GAPs (AoRga1 and AoRga2) and two PAKs (AoPak1 and AoPak2) through bioinformatics analysis and reverse genetics in Arthrobotrys oligospora, a typical nematode-trapping (NT) fungus. The transcription analyses performed at different development stages suggested that Aopaks and Aorga1 play a crucial role during sporulation and trap formation, respectively. In addition, we successfully deleted Aopak1 and Aorga1 via the homologous recombination method. The disruption of Aopak1 and Aorga1 caused a remarkable reduction in spore yield and the number of nuclei per cell, but did not affect mycelial growth. In ∆Aopak1 mutants, the trap number was decreased at 48 h after the introduction of nematodes, but nematode predatory efficiency was not affected because the extracellular proteolytic activity was increased. On the contrary, the number of traps in ∆Aorga1 mutants was significantly increased at 36 h and 48 h. In addition, Aopak1 and Aorga1 had different effects on the sensitivity to cell-wall-disturbing reagent and oxidant. A yeast two-hybrid assay revealed that AoPak1 and AoRga1 both interacted with AoRac, and AoPak1 also interacted with AoCdc42. Furthermore, the Aopaks were up-regulated in ∆Aorga1 mutants, and Aorga1 was down-regulated in ∆Aopak1 mutants. These results reveal that AoRga1 indirectly regulated AoPAKs by regulating small GTPases.
摘要：
p21-GTP酶激活的蛋白激酶（PAKs）参与RhoGTP酶下游的信号转导，受RhoGTP酶激活蛋白(Rho-GAP)调控。在这里，我们通过生物信息学分析和反向遗传学鉴定了两个直系同源Rho-GAPs（AoRga1和AoRga2）和两个PAKs（AoPak1和AoPak2）。一种典型的线虫诱捕（NT）真菌。在不同发育阶段进行的转录分析表明，Aopaks和Aorga1在孢子形成和陷阱形成中起着至关重要的作用。分别。此外,我们通过同源重组方法成功删除了Aopak1和Aorga1。Aopak1和Aorga1的破坏导致孢子产量和每个细胞的细胞核数量显着降低，但不影响菌丝生长。在ΔAopak1突变体中，引入线虫后48小时，陷阱数量减少，但线虫捕食效率不受影响，因为细胞外蛋白水解活性增加。相反,ΔAorga1突变体的陷阱数量在36h和48h时显著增加。此外，Aopak1和Aorga1对细胞壁干扰试剂和氧化剂的敏感性有不同的影响。酵母双杂交实验表明，AoPak1和AoRga1都与AoRac相互作用，AoPak1也与AoCdc42相互作用。此外，Aopaks在ΔAorga1突变体中上调，Aorga1在ΔAopak1突变体中下调。这些结果表明,AoRga1通过调节小GTP酶间接调节AoPAKs。
