CLONING AND CHARACTERIZATION OF NITRATE TRANSPORTER GENES FROM THE DIATOM CYLINDROTHECA FUSIFORMIS

Abstract

We have cloned and characterized the first nitrate transporter genes in a marine organism, from the diatom Cylindrotheca fusiformis. A cDNA clone and two genomic sequences, NAT1 and NAT2, were isolated. The sequences were highly conserved, with only 9 out of 484 amino acids differing. At least four copies of NAT genes were present in C. fusiformis, and multiple copies were present in other diatom species. NAT transcript abundance was high in cultures grown in the presence of nitrate, at nearly the same level during nitrogen starvation, and also high in urea‐grown cultures. Lower mRNA levels occurred in nitrite‐grown cultures. Transcript levels were highly repressed with NH4Cl or NH4NO3 as the nitrogen source, although very low amounts were detected. NAT mRNA levels were not directly regulated by light or dark. NAT mRNA levels varied, suggesting that nitrate uptake was not continuous, during progression through the cell cycle. The results suggest that transcriptional control contributes significantly to the overall control of nitrate uptake in C. fusiformis. The cloning of the nitrate transporter genes provides a new tool for investigating diatom nitrogen uptake and metabolism. In addition, the regulation of NAT expression by nitrogen source is likely to be useful in developing techniques to specifically control the expression of genes fused to NATregulatory sequences in transgenic diatoms.