AtAAP1/AtAAP5 Mediates The Uptake Of Amino Acid Guided Pesticides In Arabidopsis Thaliana

Amino acid-directed pesticides are synthesized by coupling pesticides with a natural amino acid as a directing group.In this thesis,functions of amino acid transporters AtAAP1 and AtAAP5 in the mediating of amino acid-directed pesticides uptake in Arabidopsis thaliana are explored.The details are as follows:(1)Root uptake experiments in the wild-type(Col-0)seedlings and plants show that,three amino acid conjugates CAP-Ala-2,Fipronil-amide-Met-2 and Fipronil-amide-Tyr,were of excellent transduction properties in the 79 chemicals tested.(2)The three genotypes of AtAAP1 or AtAAP5 seedlings and plants were applied to test their root uptake ability of CAP-Ala-2,respectively.The results show that uptake of CAP-Ala-2 in A.thaliana were increased significantly when overexpressed of AtAAP1 or AtAAP5.However,there was no significant decrease in the uptake of CAP-Ala-2 when knockout neither the AtAAP1 nor AtAAP5.These results indicated that amino acid transporters AtAAP1 and AtAAP5 were involved in the uptake of CAP-Ala-2 in A.thaliana roots,and the function between them may overlap.(3)Leaf protoplast derived from different genotypes AtAAP1 and AtAAP5 were used to test their abilities in uptake of CAP-Ala-2.The results showed that there was a high correlation between the uptake of CAP-Ala-2 and the expression level of AtAAP1 or AtAAP5.The results confirmed that the AtAAP1 and AtAAP5 participate in the process of uptake of CAP-Ala-2 in A.thaliana cells.(4)The CAP-Ala-2 was used to treat Xenopus laevis oocytes expressed the AtAAP1 or AtAAP5 and voltage-clamp was used to record changes in oocyte membrane potential.It was found that the CAP-Ala-2 failed to induce significant changes in the membrane potential of oocyte membranes expressing the AtAAP1 or AtAAP5 genes.The negative result implied that AtAAP1 and AtAAP5 transporters involved in the transport of the CAP-Ala-2 indirectly,or the electrophysiological method was not suitable for the detection of the transport of the CAP-Ala-2 by two transporters due to their transport characteristic.However,the uptake of CAP-Ala-2 in X.oocytes expressed AtAAP1 or AtAAP5 were significantly higher than that in control X.oocytes injected with water,which confirmed that AtAAP1 and AtAAP5 transported CAP-Ala-2.This dissertation demonstrated that the amino acid transporters AtAAP1 and AtAAP5 were involved in the CAP-Ala-2 uptake in Arabidopsis thaliana root.The identification of amino acid transporters functions as amino-acid-guided pesticide carriers provides the possibility to improve the bioavailability of pesticides by means of gene regulation.