| In eukaryotes,F-box protein is involved in the regulation of intracellular protein degradation,receptor recognition,signal transduction and other biological processes in the ubiquitin-proteasome pathway(UPP).They are regulatory subunits of the ubiquitin ligase complex(Skpl-Cullinl-F-box,SCF)and are involved in the specific recognition of degraded substrates.To explore the function of the F-box family gene MoFbr7 and MoFbcl in M.oryzae,MoFbr7 and MoFbcl genes were cloned from Magnaporthe oryzae and analyzed in this study.The main research results are summarized as follows:(1)Sequence analysis showed that MoFbr7 has an F-box domain at the N-terminal and a WD40 repeats at the C-terminal,and was highly conserved among filamentous fungi based on the phylogenetic analysis.Four knockout mutants of ΔMoFbr7 were obtained by gene knockout method and a complementary strain was constructed based on the deletion mutant.Phenotypic analysis showed that there were no significant differences in dry weight,sporulation,conidial production,protoplast release and pathogenicity compared with the wild-type stain Guy11.However,the growth rate ofΔMoFbr7 was significantly decreased on MM and RDC medium.The knockout strains were sensitive to the cell wall stressors Calcofluor white(CFW)and Congo Red(CR).(2)The protein domain of MoFbcl was analyzed by bioinformatics method.MoFbcl has an F-box domain at the N-terminal and 3 UIM motifs at the C-terminal,and was highly conserved among filamentous fungi based on the phylogenetic analysis.Homologous recombination techniques were used to obtain the knockout and complementary strains of MoFbcl gene.The knockout strains exhibited no difference to wild-type in dry weight,conidiation,appressorium formation,protoplast release and virulence.However,the decrease in the growth rate on MM and RDC cultures indicated that the mutant of MoFbcl gene may have abnormal utilization of some nutrients.It is sensitive to the cell wall stressors CFW and CR,suggesting that alteration of cell wall integrity of MoFbcl may occur in the ΔMoFbc1 mutants.The above results indicated that MoFbr7 and MoFbcl was involved in vegetative growth and cell wall integrity,supplying data for further revealing the regulatory mechanism of MoFbr7 and MoFbc1 in the growth and development of M.oryzae. |
