GmWRKY33A正向调控大豆抗病性

GmWRKY33A positively regulates disease resistance in soybean(Glycine max)

WRKY转录因子基因家族是植物特有的转录因子,在植物防御中起着重要作用。拟南芥中的研究表明WRKY作用于丝裂原活化蛋白激酶(mitogen activated-protein kinase,MAPK)级联途径下游,通过激活防御相关基因的表达而参与防御反应。然而大豆WRKY基因家族在防御中的作用尚不明晰。本研究通过生物信息学分析,在大豆中找到3对GmWRKY33同源基因。前2对GmWRKY33同源基因间的同源性高于84%(命名为GmWRKY33A),而每对GmWRKY33A同源基因间的同源性高于95%。从这4个GmWRKY33A同源基因保守区域选取300 bp片段构建至菜豆豆荚斑驳病毒-(bean pod mosaic virus,BPMV)改造的沉默载体(BPMV-VIGS)上以达到同时沉默上述4个GmWRKY33A基因的目的。结果表明,同时沉默上述4个GmWRKY33A基因并未改变沉默植株的发育表型,但沉默植株对大豆斑点病菌、大豆斑疹病菌和大豆花叶病毒的抗性却显著降低,说明GmWRKY33A不参与大豆的生长发育,但却参与大豆免疫反应。GmWRKY33A沉默植株中大豆斑点病菌侵染所诱导的GmMPK3和GmMPK6的激活程度显著低于空载体植株,表明GmWRKY33A可以通过调控GmMPK3/6的转录激活或激酶活性而参与大豆的免疫反应,GmWRKY33A是大豆免疫反应的正调控因子。

The WRKY transcription factor gene family is a plant-specific transcription factor that plays important roles defense responses.Studies in model plant Arabidopsis demonstrated that WRKYs function downstream of mitogen activated-protein kinase(MAPK)signaling cascade and participate in defense responses through activating the expression of defense-related genes.However,the roles of WRKYs in defense responses have not been previously investigated in paleopolyploidy soybean.Bioinfomatic analysis revealed that there are three pair of GmWRKY33 genes in the soybean genome.The identity of first two pair of GmWRKY33 genes is greater than 84%(named as GmWRKY33A).The identity of genes within the same pair is greater than 95%.A 300 bp fragment highly homologous to these four GmWRKY33A was chosen to clone into bean pod mosaic virus(BPMV)-based silencing vector(BPMV-VIGS)to achieve the goal of silencing four GmWRKY33A genes simultaneously.In this study,we simultaneously silenced four homologous genes of GmWRKY33A using a bean pod mottle virus(BPMV)vector carrying a single fragment of GmWRKY33A.Comparing the silenced plants with the vector control plants,no evident morphological phenotypes were observed.However,the GmWRKY33A-silenced plants exhibited significantly reduced resistance to Pseudomonas syringae pv.glycinea(Psg),Xanthomonas axonopodis pv.glycine(Xag),as well as to soybean mosaic virus(SMV).Furthermore,we demonstrated that silencing these GmWRKY33A genes significantly inhibited the activation of GmMPK3/GmMPK6 induced by Psg infection.Collectively,our results suggest that GmWRKY33As are involved in soybean immunity through regulating the transcription of GmMPK3/6 genes or activating the kinase activities of GmMPK3/6.Taken together,our results demonstrated that GmWRKY33As are positive regulators of soybean immune responses.