Evidence for post-translational regulation of NrtA, the Aspergillus nidulans high-affinity nitrate transporter

Here, influx and efflux of (13)NO(3)(-), and net fluxes of (14)NO(3)(-) and (14)NO(2)(-), were measured in Aspergillus nidulans mutants niaD171 and niiA5, devoid of nitrate reductase (NR) and nitrite reductase (NiR) activities, respectively. Transcript and protein abundances of NrtA, the A. nidulans principal high-affinity NO(3)(-) transporter, were determined using semiquantitative reverse transcription-polymerase chain reaction and western blots, respectively. (13)NO(3)(-) influx in niaD171 was negligible relative to wild-type values, whereas efflux to influx ratios increased nine-fold. Nevertheless, NrtA mRNA and NrtA protein were expressed at levels more than two-fold and three-fold higher, respectively, in niaD171 than in the wild-type strain. This is the first demonstration of diminished high-affinity NO(3)(-) influx associated with elevated transporter levels, providing evidence that, in addition to transcriptional regulation, control of NrtA expression operates at the post-translational level. This mechanism allows for rapid control of NO(3)(-) transport at the protein level, reduces the extent of futile cycling of NO(3)(-) that would otherwise represent a significant energy drain when influx exceeds the capacity for assimilation or storage, and may be responsible for the rapid switching between the on and off state that is associated with simultaneous provision of NH(4)(+) to mycelia absorbing NO(3)(-).