Expression And Functional Analysis Of Transcription Factor MYB53 In Arabidopsis

Developmental phase change in plants affects a plethora of important traits,such asplant productivity,biomass,reproductive competenence,pest and diseasse resistance,abiotic stress tolerance,secondary metabolite synthesis,and so on.Thereforethe studyofdevelopmental phase changehas great biological significance.miR156 has been shown to be the master regulator of vegetative phase change in plants.The function of SPLs,target of miR156,has been extensively studied,but the study ofhow miR156 is regulated by upstream factors is just emerging.miR156 expression was shown to respond to ambient temperature,sugar,CO2 and N,and some proteins in Arabidopsis including AGL15/18,AtBMI1 and PNY have been shownto regulate miR156 expression by directly binding to the promoter regions of miR156,but how miR156 responds to these externalcuesand the biological significance of this direct regulation remain elusive.In an effort to identify co-expressed genes with miR156 in a microarray data generaged using RNA from shoot apices at different developmental stages in Arabidopsis,we identified that MYB53,a R2R3 MYB transcription factor,has an analogous expression pattern with miR156,i.e.the expression level declines with time.To uncover the link between MYB53 and miR156 in Arabidopsis,we performed the following studies using genetic and molecular biology tools.Results are as follows:1.qRT-PCR confirmed that the expression of MYB53 declinesas plants age,similar to the expression of miR156.GUS reporter lines showed that the expression of MYB53 and miR156 A overlaps spatially,and,MYB53 expression exhibitsa rhythmic pattern.2.Phenotypic characterization of plants overexpressing MYB53 indicated that overexpression of MYB53 delayed flowering.The expression of miR156,SOC1,and SVP were elevated in MYB53 overexpresser,whereasthe expressions of SPL3,SPL9,and miR172 were reduced.3.Genetic analysis indicated that rSPL9 transgenic plants and toe1/2 mutants could partially recover the delayed flowering phenotype of UBI10::MYB53 plants.This result suggests that MYB53 functionsupstream of miR156 to regulate flowering in Arabidopsis.4.Computational analysis indicated that MYB53 transcripthas a potential miR167 binding site,and we verified that MYB53 mRNA was cleaved by miR167using5'RLM-RACEmethod.5.MYB53 could physically interact with BRM(BRAHMA),a chromatin remodeler,in yeast.BRM involvesphase change by regulating miR156,implying a potential role of MYB53-BRM in regulation of flowering.Our study above showed that MYB53 plays a much more important role inregulation of flowering in plant development by acting on the miR156-SPL pathway.