title: Putative RhoGAP proteins orchestrate vegetative growth, conidiogenesis and pathogenicity of the rice blast fungus Magnaporthe oryzae.
abstract: Rho GTPases, acting as molecular switches, are involved in the regulation of diverse cellular functions. Rho GTPase activating proteins (Rho GAPs) function as negative regulators of Rho GTPases and are required for a variety of signaling processes in cell development. But the mechanisms underlying Rho GAPs in Rho-mediated signaling pathways in fungi are still elusive. There are eight RhoGAP domain-containing genes annotated in the Magnaporthe oryzae genome. To understand the function of these RhoGAP genes, we generated knockout mutants of each of the RhoGAP genes through a homologous recombination-based method. Phenotypic analysis showed that growth rate of aerial hyphae of the Molrg1 deletion mutant decreased dramatically. The ΔMolrg1 mutant showed significantly reduced conidiation and appressorium formation by germ tubes. Moreover, it lost pathogenicity completely. Deletion of another Rho GAP (MoRga1) resulted in high percentage of larger or gherkin-shaped conidia and slight decrease in conidiation. Appressorial formation of the ΔMoRga1 mutant was delayed significantly on hydrophobic surface, while the development of mycelial growth and pathogenicity in plants was not affected. Confocal fluorescence microscopy imaging showed that MoRga1-GFP localizes to septal pore of the conidium, and this localization pattern requires both LIM and RhoGAP domains. Furthermore, either deleting the LIM or RhoGAP domain or introducing an inactivating R1032A mutation in the RhoGAP domain of MoRga1 caused similar defects as the Morga1 deletion mutant in terms of conidial morphology and appressorial formation, suggesting that MoRga1 is a stage-specific regulator of conidial differentiation by regulating some specific Rho GTPases. In this regard, MoRga1 and MoLrg1 physically interacted with both MoRac1-CA and MoCdc42-CA in the yeast two-hybrid and pull-down assays, suggesting that the actions of these two GAPs are involved in MoRac1 and MoCdc42 pathways. On the other hand, six other putative Rho GAPs (MoRga2 to MoRga7) were dispensable for conidiation, vegetative growth, appressorial formation and pathogenicity, suggesting that these Rho GAPs function redundantly during fungal development. Taking together, Rho GAP genes play important roles in M. oryzae development and infectious processes through coordination and modulation of Rho GTPases.
authors.affiliation: Key Laboratory of Bio-pesticide and Chemistry Biology, Ministry of Education, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, China.
authors.forename: Wenyu
authors.initials: W
authors.lastname: Ye
authors.affiliation: Key Laboratory of Bio-pesticide and Chemistry Biology, Ministry of Education, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, China.
authors.forename: Xiao
authors.initials: X
authors.lastname: Chen
authors.affiliation: Key Laboratory of Bio-pesticide and Chemistry Biology, Ministry of Education, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, China.
authors.forename: Zhenhui
authors.initials: Z
authors.lastname: Zhong
authors.affiliation: Key Laboratory of Bio-pesticide and Chemistry Biology, Ministry of Education, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, China.
authors.forename: Meilian
authors.initials: M
authors.lastname: Chen
authors.affiliation: Key Laboratory of Bio-pesticide and Chemistry Biology, Ministry of Education, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, China.
authors.forename: Lei
authors.initials: L
authors.lastname: Shi
authors.affiliation: Key Laboratory of Bio-pesticide and Chemistry Biology, Ministry of Education, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, China.
authors.forename: Huakun
authors.initials: H
authors.lastname: Zheng
authors.affiliation: Key Laboratory of Bio-pesticide and Chemistry Biology, Ministry of Education, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, China.
authors.forename: Yahong
authors.initials: Y
authors.lastname: Lin
authors.affiliation: Key Laboratory of Bio-pesticide and Chemistry Biology, Ministry of Education, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, China.
authors.forename: Dongmei
authors.initials: D
authors.lastname: Zhang
authors.affiliation: Key Laboratory of Bio-pesticide and Chemistry Biology, Ministry of Education, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, China.
authors.forename: Guodong
authors.initials: G
authors.lastname: Lu
authors.affiliation: Key Laboratory of Bio-pesticide and Chemistry Biology, Ministry of Education, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, China; Department of Biochemistry and Molecular Biology, University of Oklahoma Health Sciences Center, Oklahoma City, OK 73104, USA.
authors.forename: Guangpu
authors.initials: G
authors.lastname: Li
authors.affiliation: Key Laboratory of Bio-pesticide and Chemistry Biology, Ministry of Education, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, China. Electronic address: sishenbb@163.com.
authors.forename: Jisheng
authors.initials: J
authors.lastname: Chen
authors.affiliation: Key Laboratory of Bio-pesticide and Chemistry Biology, Ministry of Education, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, China. Electronic address: wangzh@fjau.edu.cn.
authors.forename: Zonghua
authors.initials: Z
authors.lastname: Wang
mesh_terms: D000595:Amino Acid Sequence; D002473:Cell Wall / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D020690:GTPase-Activating Proteins / Q000378:metabolism*; D017353:Gene Deletion; D020082:Magnaporthe / Q000254:growth & development / Q000378:metabolism / Q000472:pathogenicity*; D008969:Molecular Sequence Data; D009154:Mutation; D012275:Oryza / Q000382:microbiology*; D013172:Spores, Fungal / Q000254:growth & development
chemical_list: D005656:Fungal Proteins; D020690:GTPase-Activating Proteins; C085379:rho GTPase-activating protein
keywords: Appressorial formation; Conidial morphogenesis; Magnaporthe oryzae; Pathogenicity; Rho GAP; Rho GTPase