| Heterotrimeric GTP-binding proteins(G-proteins),which consist of Gα,Gβ and Gγ subunits,play important roles in transducing extracellular signals perceived by cell surface receptors into intracellular physiological responses.Gβ is encoded by a single gene AGB1 in Arabidopsis.Loss of AGB1 in Arabidopsis leads to enhanced susceptibility to almost all types of pathogens.However,the involvement of soybean AGB1 in defense responses has not been investigated previously.Soybean is a tetraploid plant,the traditional mutant-screening method is not suitable for studying the gene function of soybean.In soybean(Glycine max),virus-induced gene silencing(VIGS)mediated by Bean pod mottle virus(BPMV)has been used successfully to silence soybean genes and overcome the problem of gene redundancy.Through bioinformatics analysis,we identified 4 GmAGB1 homologous genes in soybean genome sharing homology of 86-97%.A 354-bp fragment,which is conserved among these four GmAGB1 homologs and expected to silence these four genes simultaneously,was cloned into the BPMV vector.The VIGS construct was delivered into soybean plants by bolisctic barbombardment.Silencing GmAGB1 homologs in soybean resulted in a dwarfed phenotype,suggesting a role of GmAGB1 in growth and development in soybean.Pathogen inoculation assay on GmAGB1-silenced plants indictated that silencing GmAGB1 genes reduced the resistance of soybean against Xanthomonas campestris pv.glycines(Xag).This results indicated that GmAGB1 gene plays a positive regulatory role in Xag resistance.Production and over-accumulation of reactive oxygen species(ROS)in response to pathogen infections is one of strageties plants used to defend pathogens.It was found that the accumulation of ROS induced by Pseudomonas syringae pv.glycinea(Psg)infection on the leaves of the GmAGB1-silenced plants was significantly lower than on the leaves of the empty vector control plants,indicating that GmAGB1 is indispensable for the ROS production induced by pathogen infection.The decreased disease resistance of the GmAGB1-silenced plants may be related to the decrease of ROS accumulation induced by bacterial infection.No significant difference in the activation of Gm MPK3/6 kinase activity induced by flg22,a conserved N-terminal peptide of flagellin protein,was observed between the GmAGB1-silenced plants and vector control plants,suggsesting that the GmAGB1 is not required for activation of Gm MPK3/6.Yeast two hybrid experiment showed that GmAGB1 interacted with Gm AGG1,implying that GmAGB1 and Gm AGG1 function in a Heterotrimeric G protein complex in soybean. |
