Influences of nitrogen deficiency on the transcript levels of ammonium transporter, nitrate transporter and glutamine synthetase genes in Isochrysis galbana (Isochrysidales, Haptophyta)

Abstract

Three DNA fragments belonging to genes involved in nitrogen uptake and assimilation, including a nitrate transporter gene (IgNrt2), an ammonium transporter gene (IgAmt), and a glutamine synthetase gene (IgglnII, were obtained from the marine haptophyte, Isochrysis galbana. Based on alignments of the deduced amino acid sequences, the IgNRT2 fragment shared 47%,, identity with the Cylindrotheca fusiformis (Bacillariophyceae) nitrate transporter, the IgAMT fragment shared 48%, identity with the C. fusiformis ammonium transporter, and the IgGSII fragment shared 61% identity with Skeletonema costatum (Bacillariophyceae) glutamine synthetase. Southern hybridization indicated that both IgNrt2 and IgAmt may have more than one copy in the genome of L galbana, but only a single copy was detected for IgglnII. The transcript abundances of these three genes under various nitrogen-supply conditions were monitored simultaneously by quantitative real-time polymerase chain reaction (PCR). In the presence of ammonium, both IgNrt2 and IgAmt were severely repressed with mean mRNA concentrations of 0.67 and 3.45 mu mol (mol 18S rRNA)(-1). IgNrt2 and IgAmt mRNA transcripts increased 41-fold and 8.3-fold, respectively, in the presence of nitrate, and increased 160-fold and 15-fold, respectively, in the ni trogen-depleted condition. The mRNA level of IgglnII was low in cells grown in the presence of ammonium [3.98 mu mol (mol 18S rRNA)(-1)], but mild increases in the range of 2- to 4-fold were observed in cells grown in both nitrate-containing and nitrogen-depleted cultures. Our results demonstrated that the expression patterns of IgNrt2, IgAmt, and IgglnII shared a common trend with their homologous genes in diatoms and green algae under various nitrogen-supply conditions. This group of genes could be used as indicators in natural environments to estimate the degree of nitrogen deficiency and chemical forms of nitrogenous nutrients in use on a species-specific basis.