Nitric oxide inhibits transcription of the Na+-K+-ATPase alpha 1-subunit gene in an MTAL cell line

Nitric oxide (NO) has been implicated as an autocrine modulator of active s odium transport;. To determine whether tonic exposure to NO influences acti ve sodium transport in epithelial cells, we established transfected medulla ry thick ascending limb of Henle (MTAL) cell lines that overexpressed NO sy nthase-2 (NOS2) and analyzed the effects of deficient or continuous NO prod uction [with or without N-G-nitro-L-arginine methyl ester (L-NAME) in the c ulture medium, respectively] on Na+-K+-ATPase function and expression. The NOS2-transfected cells exhibited high-level NOS2 expression and NO generati on, which did not affect cell viability or cloning efficiency. NOS2-transfe cted cells were grown in the presence of vehicle, N-G-nitro-D-arginine meth yl ester (D-NAME), or L-NAME for 16 h, after which Rb-86(+) uptake assays, Northern analysis, or nuclear run-on transcription assays were performed. T he NOS2-transfected cells allowed to produce NO continuously (vehicle or D- NAME) exhibited lower rates of ouabain-sensitive Rb-86(+) uptake (similar t o 65%), lower levels of Na+-K+-ATPase alpha 1-subunit mRNA(similar to 60%), and reduced rates of de novo Na+-K+-ATPase al-subunit transcription compar ed with L-NAME-treated cells. These results have uncovered a novel effect o f NO to inhibit transcription of the Na+-K+-ATPase alpha 1-subunit gene.