Functional Analysis Of Cytokinin Transporter AtABCG14 In The Aerial Tissues Of Arabidopsis

Cytokinin is an important phytohormone which regulates plant growth and development,and has close relationship with leaf senescence,plant disease resistance and abiotic stresses.In contrast to its synthesis and signal transduction,the molecular mechanism of cytokinin transport is poorly understood.AtABCG14 and AtPUP14 are two types cytokinin transporters which were recently reported in plant.AtABCG14 is mainly responsible for the long-distance translocation of cytokinins,while AtPUP14 is responsible for intracellular transport of cytokinins.AtABCG14 is the first transporter involved in long-distance transport of cytokinins,which regulates the acropetal translocation of root-synthesized cytokinins in plants.AtABCG14 gene was strongly expressed in the roots of Arabidopsis thaliana,and the expression of AtABCG14 was also observed in the aerial tissues(mainly in the vein and inflorescence).Therefore,we gave a hypothesis that AtABCG14 may have some biological function in the aerial tissues.In this study,we combined the technologies such as grafting,GUS assays and isotope tracing involved in genetics,molecular biology,developtenal biology.We have drawn conclusitons listed as below:(1)When the abcg14 mutant was grafted as scion and the wild-type as rootstock,the phenotype of aerial tissue was not restored,indicating that AtABCG14 gene has biological functions in aerial tissues.(2)The relative expression of the cytokinin response genes and the expression level of ARR5::GUS in the aerial tissues of the grafts were detected in the grafted plants.The results showed that the expression level of cytokinin response gene significantly reduced in the aerial tissues of grafts in which abcgl4 mutant was used as scion and Col-4 as rootstock.It was further suggested that AtABCG14 has biological function in the aerial tissues.(3)The tissue specific expression of AtABCG14 in the aerial tissue was proved to in the phloem region by GUS assays and complementation experiments of abcgl4 mutants with vairous promoter-fused AtABCG14 gene.(4)The C13-tZ and C14-tZ exogenous absorption assays were used to detect the isotope-labeled cytokinin contents in the aerial grafted tissues.The results show that cytokinin can be transported through the vascular organization to the aerial tissue in all grafted plants.(5)The fluorescence intensity in the roots of a grafted plant with ARR5::eGFP/WT as a rootstock and ARR5::eFP/abcg14 as a scion was enhanced,showing that the translocation of cytokinin was inhibited when the AtABCG14 was knock-out in the aerial tissues.In summary,this study demonstrated that AtABCG14 participated in cytokinin distribution in Arabidopsis thaliana.In addition,when the root-synthesized cytokinins in the aerial tissues can not be successfully distributed,the cytokinins will accumulate in the roots,suggesting that loss of AtABCG14 in the aerial tissues may effect the translocation of cytokinin from roots to the shoots.