The Preliminary Function Analysis Of Arabidopsis Thaliana Magnesium Transport 4

AtMGT4 is one of the magnesium transporter gene family members with an open reading frame containing 1455 base pairs, and codes a peptide of 484 amino acid residues.The previous data in our lab showed AtMGT4 can complement the defect in magnesium-uptake of MM281 mutant strains, suggesting the role of AtMGT4 as a Mg2+ transporter. To date, however, little is known about the in vivo function(s) that AtMGT4 played in Arabidopsis. In this research, we conducted preliminary analyses on the AtMGT4 gene expression pattern, subcellular localization, and magnesium uptake activity in Arabidopsis etc, and the results showed as below.GFP-AtMGT4 was transformed into the mesophyll protoplasts of Arabidopsis, and the GFP fluorescent signal was detected in the nuclei under the fluorescence microscopy, indicating AtMGT4 protein was localized in the nucleus of a cell.The AtMGT4pro:GUS transgenic plants were obtained, and the GUS signals were found mainly in root tips, the transition region between shoot and root, the leaf apex, anthers and ovules, suggesting AtMGT4 was expressed in the above tissues.Two homozygous AtMGT4 T-DNA mutants(GK-142H10 and GK-366F02) were identified, and their T-DNA.insert positions were also determined by sequencing. In GK-142H10, the insert positions were located at -55 bp and -92 bp upstream from AtMGT4 initiation code ATG. Meanwhile, it was located at -93 bp upstream from AtMGT4 initiation code ATG in GK-366F02. The AtMGT4 transcripts displayed no obvious changes in GK-142H10 and increases a lot in GK-366F02 by semi-quantitative RT-PCR, indicating they were not valid T-DNA mutants, and the GK-366F02 mutant plant was substantially a AtMGT4 overexpression plant.We also constructed 35S:AtMGT4 transgenic plants MGT4-ox. Under low magnesium condition (10 μM Mg2+), both MGT4-ox and GK-366F02 plants displayed growth retardation, shorter main root and less lateral roots phenotype, compared to wild type plants.We further measured the magnesium concentration of the whole plants, roots and shoot tissues of GK-366F02 plants under low magnesium condition by using FAAS. The magnesium concentration of the GK-366F02 whole plants was lower than the level of the wild type, whereas the magnesium concentration in roots was stable between GK-366F02 and WT, however, the magnesium concentration in the shoot tissues of GK-366F02 was significantly lower than in WT.By transplanting the GK-366F02 from 1 mM Mg2+ hydroponic liquid to 10 μM Mg2+ hydroponic liquid, the plants were induced by the low Mg2+ stress, the AtMGTl、AtMGT2、AtMGT3、AtMGT6 and AtMGT10 genes transcripts in GK-366F02 decreased significantly after 2 days.A yeast two-hybrid(Y2H) cDNA library was screened by using AtMGT4 as the "bait" and found several peptide sequences.After the self-activation detection of these candidates, two proteins interacting with AtMGT4 protein were indentified preliminarily, they were Atlg61170 and At5g13980.Atlg61170 is an unknown protein, predicted subcellular localization of which is in the nucleus of a cell. At5g13980 is one of Glycosyl hydrolase family members.To sum up, this paper suggested in a low magnesium condition overexpression of AtMGT4 can influenced transcription of AtMGT1、 AtMGT2、AtMGT3、AtMGT6 and AtMGT10, also influenced the Mg2+ transport from roots to leaf.