Heterologous functional analysis of the Malus xiaojinensis               MxIRT1 gene and the His-box motif by expression in yeast.
Malus xiaojinensis is an important, iron-efficient rootstock germplasm. Iron uptake is an elaborately controlled process in plant roots, involving specialized transporters. MxIRT1, a Fe(II) transporter gene of M. xiaojinensis, is homologous to other iron transporters at the amino acid level. In the current study, the plasmid pYES2.0-MxIRT1, containing MxIRT1 cDNA, was constructed and transformed into yeast mutants. The results indicated that it could reverse the phenotype of yeast strain DEY1453, an iron uptake mutant. Complementation tests suggested that it might not be a specific transporter, as it was able to restore the phenotypes of other yeast mutant strains, including Mn, Cu and Zn uptake mutants. The functions of the critical histidine residues in the His-box of MxIRT1 were tested by transforming mutant yeast strain DEY1453 with different His residues altered by directed mutagenesis. The His-box of MxIRT1 was found to be necessary for iron transport, with different histidine residues (H(1-4)) playing different roles in the transport.
Institute for Horticultural Plants, China Agricultural University, 2 Yuanmingyuan West Road, Haidian District, Beijing, 100193, People's Republic of China.
Xue-Ning
XN
Zhang
Zhen-Hai
ZH
Han
Li-Li
LL
Yin
Jin
J
Kong
Xue-Feng
XF
Xu
Xin-Zhong
XZ
Zhang
Yi
Y
Wang
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D027682:Cation Transport Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000502:physiology; D007501:Iron / Q000378:metabolism; D027845:Malus / Q000235:genetics*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010940:Plant Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000502:physiology; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000254:growth & development / Q000378:metabolism
D027682:Cation Transport Proteins; D010940:Plant Proteins; D011994:Recombinant Proteins; D007501:Iron