Molecular cloning and identification of an ammonium transporter gene from pear

The molecular basis of ammonium acquisition in fruit trees remains poorly understood. Here, a new ammonium transporter gene, PbAMT1;1, was isolated from Pyrus betulifolia. The full length of PbAMT1;1 was 1,693 bp, coding a protein of 505 amino acids. The deduced structure of the PbAMT1;1 protein revealed 11 membrane-spanning domains and an ammonium transporter signature motif 'D(204)FAGSGMVHMVGGIAGLWGAFIESPR(229)' located in the fifth predicted transmembrane region. PbAMT1;1 transcripts were detectable in all parts of the seedlings and predominantly located to the roots. In addition, a change of pH had little effect on its expression patterns. PbAMT1;1 expression levels in roots were strongly induced by ammonium depletion and suppressed by high ammonium. However, its transcription levels in leaves were reduced by free nitrogen and increased after high amounts of ammonium were supplying. Further, PbAMT1;1 exhibited diurnal rhythms with the highest expression at noon. When abscisic acid and methyl jasmonate existed, the mRNA abundance of PbAMT1;1 increased. The results of yeast complementation showed that the PbAMT1;1 gene could help yeast mutant stain 31019b to grow in a media with micromole ammonium concentrations. Furthermore, PbAMT1;1 mediated NH4 (+) uptake with high affinity producing K (m) values from 20.9 to 23.2 mu M at different pH values. However, the net (NH4)-N-15 (+) uptake K (m) of PbAMT1;1 showed no significant differences at pH 4.8, 5.8 or 6.8. In conclusion, PbAMT1;1 is a functional AMT gene having the features of the AMT1 family. Its expression was regulated by ammonium concentration, diurnal rhythms and plant hormones, but not by external pH levels.