Ethylene treatment promotes orange coloration in the flavedo of Satsuma mandarin(Citrus unshiu Marc.)fruit,but the corresponding regulatory mechanism is still largely unknown.In this study,we identified a C2H2-type zi...Ethylene treatment promotes orange coloration in the flavedo of Satsuma mandarin(Citrus unshiu Marc.)fruit,but the corresponding regulatory mechanism is still largely unknown.In this study,we identified a C2H2-type zinc-finger transcription factor,CitZAT4,the expression of which was markedly induced by ethylene.CitZAT4 directly binds to the Cit PSY promoter and activates its expression,thereby promoting carotenoid biosynthesis.Transient expression in Satsuma mandarin fruit and stable transformation of citrus calli showed that overexpressing of CitZAT4 inhibited Cit LCYE expression,thus inhibitingα-branch yellow carotenoid(lutein)biosynthesis.CitZAT4 overexpression also enhanced the transcript levels of CitLCYB,Cit HYD,and CitNCED2,promotingβ-branch orange carotenoid accumulation.Molecular biochemical assays,including yeast one-hybrid(Y1H),electrophoretic mobility shift(EMSA),chromatin immunoprecipitation quantitative polymerase chain reaction(Ch IP-q PCR),and luciferase(LUC)assays,demonstrated that CitZAT4 directly binds to the promoters of its target genes and regulates their expression.An ethylene response factor,CitERF061,which is induced by ethylene signaling,was found to directly bound to the CitZAT4 promoter and induced its expression,thus positively regulating CitZAT4-mediated orange coloration in citrus fruit.Together,our findings reveal that a CitZAT4-mediated transcriptional cascade is driven by ethylene via CitERF061,linking ethylene signaling to carotenoid metabolism in promoting orange coloration in the flavedo of Satsuma mandarin fruit.The molecular regulatory mechanism revealed here represents a significant step toward developing strategies for improving the quality and economic efficiency of citrus crops.展开更多
基金supported by the National Key R&D Program of China(2023YFD2300600)the National Natural Science Foundation of China(No.31930095)the National Modern Agricultural(Citrus)Technology Systems of China(No.CARS-27)。
文摘Ethylene treatment promotes orange coloration in the flavedo of Satsuma mandarin(Citrus unshiu Marc.)fruit,but the corresponding regulatory mechanism is still largely unknown.In this study,we identified a C2H2-type zinc-finger transcription factor,CitZAT4,the expression of which was markedly induced by ethylene.CitZAT4 directly binds to the Cit PSY promoter and activates its expression,thereby promoting carotenoid biosynthesis.Transient expression in Satsuma mandarin fruit and stable transformation of citrus calli showed that overexpressing of CitZAT4 inhibited Cit LCYE expression,thus inhibitingα-branch yellow carotenoid(lutein)biosynthesis.CitZAT4 overexpression also enhanced the transcript levels of CitLCYB,Cit HYD,and CitNCED2,promotingβ-branch orange carotenoid accumulation.Molecular biochemical assays,including yeast one-hybrid(Y1H),electrophoretic mobility shift(EMSA),chromatin immunoprecipitation quantitative polymerase chain reaction(Ch IP-q PCR),and luciferase(LUC)assays,demonstrated that CitZAT4 directly binds to the promoters of its target genes and regulates their expression.An ethylene response factor,CitERF061,which is induced by ethylene signaling,was found to directly bound to the CitZAT4 promoter and induced its expression,thus positively regulating CitZAT4-mediated orange coloration in citrus fruit.Together,our findings reveal that a CitZAT4-mediated transcriptional cascade is driven by ethylene via CitERF061,linking ethylene signaling to carotenoid metabolism in promoting orange coloration in the flavedo of Satsuma mandarin fruit.The molecular regulatory mechanism revealed here represents a significant step toward developing strategies for improving the quality and economic efficiency of citrus crops.
