Nitric oxide inhibits transcription of the Na+-K+-ATPase α1-subunit gene in an MTAL cell line

Nitric oxide (NO) has been implicated as an autocrine modulator of active sodium transport. To determine whether tonic exposure to NO influences active sodium transport in epithelial cells, we established transfected medullary thick ascending limb of Henle (MTAL) cell lines that overexpressed NO synthase-2 (NOS2) and analyzed the effects of deficient or continuous NO production [with or without N G-nitro-l-arginine methyl ester (l-NAME) in the culture medium, respectively] on Na+-K+-ATPase function and expression. The NOS2-transfected cells exhibited high-level NOS2 expression and NO generation, which did not affect cell viability or cloning efficiency. NOS2-transfected cells were grown in the presence of vehicle, N G-nitro-d-arginine methyl ester (d-NAME), orl-NAME for 16 h, after which86Rb+uptake assays, Northern analysis, or nuclear run-on transcription assays were performed. The NOS2-transfected cells allowed to produce NO continuously (vehicle ord-NAME) exhibited lower rates of ouabain-sensitive86Rb+uptake (∼65%), lower levels of Na+-K+-ATPase α1-subunit mRNA (∼60%), and reduced rates of de novo Na+-K+-ATPase α1-subunit transcription compared withl-NAME-treated cells. These results have uncovered a novel effect of NO to inhibit transcription of the Na+-K+-ATPase α1-subunit gene.