Identification Of The Activity Of Three ABA Receptor PYR/PYLs/RCAR In Zea Mays L.

Plant hormone abscisic acid(ABA)regulates a lot of plant growth and development processes and plant responses to environmental stresses,which is through the core ABA signaling way PYR/PYLs/RCAR-PP2Cs-SnRK2 s.In this core signaling pathway,ABA receptor PYR/PYLs/RCAR inhibit the protein phosphatase activity of clade A PP2 Cs,thus mediate the ABA signal.Recently,multiple studies have shown that over expression of ABA receptors could promote plant drought resistance and even improve plant water use efficiency.In previous researches,our lab found that the expression of PYR/PYLs/RCAR genes show tissue specificity and subfamily specificity,and three representative genes ZmPYL1(KJ700410.1),ZmPYL4(KJ191278.1)and ZmPYL10(KP708579)were cloned.In this study,we verified the ABA receptor activity of Zm PYL1/4/10.Furthermore,we analyzed the transcriptional responses of ZmPYL1/4/10 under ABA treatment and various stresses in maize seedings.The results are as follows:(1)The ABA receptor activity of ZmPYL1/4/10 were confirmed through in vitro phosphatase activity assay.ZmPYL1 and ZmPYL10 inhibit the protein phosphatase activity of ZmPP2Ca/ZmHAI1 in an ABA dependent way,whereas ZmPYL4 inhibit the protein phosphatase activity of ZmPP2Ca/ZmHAI1 in an ABA independent way.Both the three receptors can significantly inhibit the protein phosphatase activity of ZmPP2Ca/ZmHAI1 under 0.5?M ABA treatment,thus show their high sensitivity to ABA,which indicates their capability in improving plants stress resistance.(2)The gene expression of ZmPYL1/4/10 under ABA treatment and several abiotic stresses was analyzed by real-time fluorescent quantitative PCR.The gene expression of ZmPYL1 in maize seeding roots was upregulated by 10 times than basal level under 5mM H2O2 treatment,which show its high sensitivity to ROS,whereas ZmPYL4 was downregulated to only 10% of basal level under 10?M ABA treatment,thus show its high sensitivity to ABA.The results confirm that the tissue specificity and subfamily specificity of the transcriptional responses of PYR/PYLs/RCAR genes are ubiquitous.(3)Agrobacterium-mediated transformation of Arabidopsis thaliana was carried out using the floral dip method,and the 35s::ZmPYL1/4/10 over expression transgenic plants were obtained,which provides materials for further study of the biological functions of ZmPYL1/4/10.Further more,we found that ABA receptors ZmPYL1 and ZmPYL10 were localized in cell nucleus in transgenic plants(35s::YFP-ZmPYL1 and 35s::YFP-ZmPYL10).