The naturally selected extreme traits of zinc and cadmium hyperaccumulation and hypertolerance in Arabidopsis halleri depend on strongly elevated HEAVY METAL ATPase 4(HMA4)transcript levels compared to those in the cl...The naturally selected extreme traits of zinc and cadmium hyperaccumulation and hypertolerance in Arabidopsis halleri depend on strongly elevated HEAVY METAL ATPase 4(HMA4)transcript levels compared to those in the closely related Arabidopsis thaliana.This difference is regulated in cis;AhHMA4 upstream sequences alone are sufficient to confer increased expression,as previously demonstrated using reporter gene fusions stably introduced into both A.halleri and A.thaliana.However,the underlying cis-regulatory divergence specific to A.halleri remains unknown.Here,we identify cis-regulatory metal hyperaccumulation elements(MHEs)that increase AhHMA4 promoter activities by examining stably transformed reporter lines carrying partial deletions or mutations in AhHMA4 upstream sequences.MHE1(consensus TGTAAC)functions in the distal regions of AhHMA4 promoters,and all three tandem AhHMA4 gene copies share a proximal upstream pair of MHE2 motifs(consensus AAATATCT),corresponding to the evening element.The evening element is a known target of Arabidopsis CIRCADIAN CLOCK-ASSOCIATED 1(CCA1),a core circadian clock transcription factor that mediates light-dependent and circadian gene expression.We show that the elevated activity of the AhHMA4-1 promoter depends on MHE2 in cis and CCA1 in trans,and it can be recapitulated by introducing an intact pair of MHE2 motifs into the A.thaliana HMA4 promoter using site-directed mutagenesis.We also found that HMA4 transcript levels show diel rhythmicity in A.halleri but not in A.thaliana.In summary,we identify the causal cis-regulatory elements that co-opt a known regulator of diel and seasonal transcriptional rhythms to mediate enhanced expression of a gene critical for a naturally selected extreme trait syndrome.展开更多
基金funded by the National Council of Humanities,Sciences and Technologies(Consejo Nacional de Humanidades,Ciencias y Tecnologias[CONACYT])scholarship no.438349a scholarship for the completion of dissertations at Ruhr University Bochum from the Wilhelm and Gunter Esser Foundation(L.C.)+4 种基金by the German Research Foundation-(Deutsche Forschungsgemeinschaft[DFG])project no.5453631European Research Council(ERC)-AdG 788380"LEAP EXTREME"(U.K.)the Fonds de la Recherche Scientifique-FNRS(FRFC-2.4583.08 and PDR-T.0206.13)the University of Liege(SFRD-12/03)the Belgian Program on Interuniversity Attraction Poles(IAP no.P7/44)(M.H.).
文摘The naturally selected extreme traits of zinc and cadmium hyperaccumulation and hypertolerance in Arabidopsis halleri depend on strongly elevated HEAVY METAL ATPase 4(HMA4)transcript levels compared to those in the closely related Arabidopsis thaliana.This difference is regulated in cis;AhHMA4 upstream sequences alone are sufficient to confer increased expression,as previously demonstrated using reporter gene fusions stably introduced into both A.halleri and A.thaliana.However,the underlying cis-regulatory divergence specific to A.halleri remains unknown.Here,we identify cis-regulatory metal hyperaccumulation elements(MHEs)that increase AhHMA4 promoter activities by examining stably transformed reporter lines carrying partial deletions or mutations in AhHMA4 upstream sequences.MHE1(consensus TGTAAC)functions in the distal regions of AhHMA4 promoters,and all three tandem AhHMA4 gene copies share a proximal upstream pair of MHE2 motifs(consensus AAATATCT),corresponding to the evening element.The evening element is a known target of Arabidopsis CIRCADIAN CLOCK-ASSOCIATED 1(CCA1),a core circadian clock transcription factor that mediates light-dependent and circadian gene expression.We show that the elevated activity of the AhHMA4-1 promoter depends on MHE2 in cis and CCA1 in trans,and it can be recapitulated by introducing an intact pair of MHE2 motifs into the A.thaliana HMA4 promoter using site-directed mutagenesis.We also found that HMA4 transcript levels show diel rhythmicity in A.halleri but not in A.thaliana.In summary,we identify the causal cis-regulatory elements that co-opt a known regulator of diel and seasonal transcriptional rhythms to mediate enhanced expression of a gene critical for a naturally selected extreme trait syndrome.
