Regulation of high-affinity nitrate transporter genes and high-affinity nitrate influx by nitrogen pools in roots of barley

To investigate the regulation of HvNRT2, genes that encode high-affinity NO 3- transporters in barley (Hordeum vulgare) roots, seedlings were treated w ith 10 mM NO3- in the presence or absence of amino acids (aspartate, aspara gine, glutamate [Glu], and glutamine [Gln]), NH4+, and/or inhibitors of N a ssimilation. Although all amino acids decreased high-affinity (NO3-)-N-13 i nflux and HvNRT2 transcript abundance, there was substantial interconversio n of administered amino acids, making it impossible to determine which amin o acid(s) were responsible for the observed effects. To clarify the role of individual amino acids, plants were separately treated with tungstate, met hionine sulfoximine, or azaserine (inhibitors of nitrate reductase, Gln syn thetase, and Glu synthase, respectively). Tungstate increased the HvNRT2 tr anscript by 20% to 30% and decreased NO3- influx by 50%, indicating that NO 3- itself does not regulate transcript abundance, but may exert post-transc riptional effects. Experiments with methionine sulfoximine suggested that N H4+ may down-regulate HvNRT2 gene expression and high-affinity NO3- influx by effects operating at the transcriptional and post-transcriptional levels . Azaserine decreased HvNRT2 transcript levels and NO3- influx by 97% and 9 5%, respectively, while decreasing Glu and increasing Gin levels. This sugg ests that Gln (and not Glu) is responsible for down regulating HvNRT2 expre ssion, although it does not preclude a contributory effect of other amino a cids.