The Molecular Mechanism Of Coordinate Regulation Gene AtSRG1 Expression By AtMYC2 And AtHsf3

Heat shock factors(Hsf)play an important role in thermo-tolerance by regulating the heat shock proteins(HSPs)and heat-inducible gene Galactinol synthasel(SRG1).However,the regulatory mechanism of Hsf3 interacting with MYC2 under jasmonate and heat shock stress remains unclear.Here,we revealed that Hsf3 interacts with the AtMYC2 coordinately modulating SRG1 gene expression under synergistic jasmonate and heat shock stress.(1)AtSRG1 expression was significantly induced by jasmonate(JA),heat shock stress(HS),and JA+HS.Both qPCR and GUS assay indicated that AtSRGl expression was not only remarkably induced by HS,but also by HS,JA+HS.(2)AtMYC2 regulated gene AtSRGl expression by binding the promoter of AtSRGl,the results of EMSA showed that transcription factor AtMYC2 could specifically bind the G-box in the promoter of the AtSRG1,Meanwhile,transcripts of the gene AtSRG1 in the mutant myc2/myc3/myc4 under JA,HS,JA+HS stresses indicated that the relative expression level of AtSRG1 was significantly reduced compared to the wild type.Thus,we proposed that AtMYC2 regulated the AtSRG1 expression.(3)AtMYC2 interacted with AtHsf3,Yeast two hybrid(Y2H)assays confirmed the interaction of AtMYC2 and AtHsf3,furthermore,transcription AtHsf3 had transcription self-activation domain which located at AA410-459,and the interaction domain located at AA 410-459.(4)The mutant myc2/myc3/myc4/hsf3 homozygote were cultivated and identified,there was no difference between the fourth mutant and wild type,besides,the number of seeds in the seedpod were significantly less than the wild type and the third mutant myc2/3/4.(5)The expression AtSRG1 in the mutants hsf3,myc2/3/4,myc2/3/4/hsf3 under JA,HS,JA+HS were analyzed respectively by qPCR,the results indicated that,AtMYC2 regulated the AtSRG1 expression through JA signal pathway,Transcription including AtHsf3 regulated the AtSRGl expression under heat shock stress,AtHSF3 and AtMYC2 coordinately regulate AtSRGl gene expression under JA and HS stresses.Above all,In this study,AtSRG1 expression was significantly induced by jasmonate(JA),heat shock stress(HS),and JA+HS,EMSA assay showed AtMYC2 regulated gene AtSRG1 expression by binding the promoter of AtSRG1,,Yeast two hybrid(Y2H)assays confirmed the interaction of AtMYC2 and AtHsf3,RNA-seq and qPCR combined indicated that AtHSF3 and AtMYC2 coordinately regulate AtSRGl gene expression.