The Functional Analysis Of AtMGT3, A Member Of Putative Mg²⁺ Transporter Family In Arabidopsis

AtMGT gene family was identified by Li et al.(2001),which contains 10-members designated as AtMGT1-AtMGT10.AtMGT genes encodes putative Mg²⁺transporters,which might play an essential role in maintaining Mg²⁺homeostasis of the internal environment in the cells.AtMGT3 locates on the second chromosome,encoding a 421aa residues protein.Structural analysis reveals this protein has the common characteristics of the AtMGTs,e.g.,containing two hydrophobic transmembrane (TM)domains,and a GMN conserved motif in the first transmembrane domain.For the time being,4-members of AtMGT were found to functionally complement the bacterial or yeast Mg²⁺- deficient mutants,suggesting a potential role in Mg²⁺ transport in Arabidopsis.This study shows some results of AtMGT3 gene cloning and characterization.1.The cDNA cloning was obtained by RT-PCR,and then sequenced.It showed the identical sequence to that of the predicted by AGI.Then cDNA was subcloned to pTrc99A and transformed to the Salmonella typhimurium mutant MM281 which lack Mg²⁺transport capability and cannot survive in the medium with the Mg²⁺concentration below 10mmol/L.The results showed that AtMGT3 can functionally complement the bacterial mutant.2.The results of functional complementation and liquid growth curve analysis showed that AtMGT3 mediated low-affinity transport of Mg²⁺.Furthermore,AtMGT3 might transport other divalent cations like Fe²⁺.But,the concentration required for transport of Fe²⁺is beyond normal physiological range.Under normal physiological ranges,the major physiological function of AtMGT3 is Mg²⁺transport.3.Under the normal and lacking Mg²⁺physiological ranges,compared to the wild type,SOD activity be higher and chlorophyâ…¡concentration be lower in the mutant,we can find that AtMGT3 and Mg²⁺are related.It indirect improved that AtMGT3 can transport Mg²⁺.4.The expression pattern indicated that:AtMGT3 transcripts were expressed in roots,stems,leaves,flowers, and siliques,but exist most abundantly in flowers,least abundantly in roots.5.Meanwhile,the transgenic plants transformed with the AtMGT3::GFP fusion constructs were detected with the confocal fluorescence microscope.The result indicated that AtMGT3 locates in the plasma membrane.6.GUS expression of AtMGT3 mainly in anthers,roots and leaves,while the expression of anthers and roots have the specificity of times.7.T-DNA insertional AtMGT3 gene knock-out mutant plants and AtMGT3 overexpression transgenic plants were screened and their phenotype was analyzed.Unfortunately,there is no obvious phenotype found.