NRT1.1 and NIA1 genes, which encode a nitrate (NO3-) transporter and the minor isoform of NO3- reductase (NR), respectively, are overexpressed in roots of NR-deficient mutants of Arabidopsis grown on nutrient solution containing NO3- and reduced N. The overexpression is found only in mutants with reduced NIA2 activity, and disruption of the NIA1 gene alone has no effect on NRT1.1 expression. Because the up-regulation of NRT1.1 and NIA1 is observed in N-sufficient NR mutant plants, it cannot be related to a release of the general feedback repression exerted by the N status of the plant. Our data do not support the hypothesis of overinduction of these genes by an increased concentration of NO3- in tissues. Furthermore, although a control by external pH might contribute to the regulation of NRT1.1, changes in external pH due to lack of NR activity cannot alone explain the up-regulation of both genes. The stimulation of NRT1.1 and NIA1 in NR mutants in these conditions suggests that NR activity is able to repress directly the expression of both genes independently of the availability of reduced N metabolites in wild-type plants. Accordingly, nitrite (NO2-) strongly represses NRT1.1 and NIA1 transcript accumulation in the roots. This effect is rapid, specific, and reversible. Furthermore, transport studies on plants exposed to NO2- show that down-regulation of the NRT1.1 gene is associated with a decrease in NO3- influx. These results indicate that feedback regulation of genes of NO3- assimilation relies not only on the repression exerted by reduced N metabolites, such as NH4+ or amino acids, but may also involve the action of NO2- as a regulatory signal.