Transforming growth factor-β downregulates sGC subunit expression in pulmonary artery smooth muscle cells via MEK and ERK signaling

TGF beta activation during newborn lung injury decreases the expression of pulmonary artery smooth muscle cell (PASMC)-soluble guanylate cyclase (sGC), a critical mediator of nitric oxide signaling. Using a rat PASMC line (CS54 cells), we determined how TGF beta downregulates sGC expression. We found that TGF beta decreases sGC expression through stimulating its type I receptor; TGF beta type I receptor (TGF beta R1) inhibitors prevented TGF beta-1-mediated decrease in sGC beta 1 subunit mRNA levels in the cells. However, TGF beta R1-Smad mechanisms do not regulate sGC; effective knockdown of Smad2 and Smad3 expression and function did not protect sGC alpha 1 mRNA levels during TGF beta-1 exposure. A targeted small-molecule kinase inhibitor screen suggested that MEK signaling regulates sGC expression in TGF beta-stimulated PASMC. TGF beta activates PASMC MEK/ERK signaling; CS54 cell treatment with TGF beta-1 increased MEK and ERK phosphorylation in a biphasic, time- and dose-dependent manner. Moreover, MEK/ERK activity appears to be required for TGF beta-mediated sGC expression inhibition in PASMC; MEK and ERK inhibitors protected sGC beta 1 mRNA expression in TGF beta-1-treated CS54 cells. Nuclear ERK activity is sufficient for sGC regulation; heterologous expression of a nucleusretained, constitutively active ERK2-MEK1 fusion protein decreased CS54 cell sGC alpha 1 mRNA levels. The in vivo relevance of this TGF beta-MEK/ERK-sGC downregulation pathway is suggested by the detection of ERK activation and sGC alpha 1 protein expression downregulation in TGF beta-associated mouse pup hyperoxic lung injury, and the determination that ERK decreases sGC alpha 1 protein expression in TGF beta-1-treated primary PASMC obtained from mouse pups. These studies identify MEK/ERK signaling as an important pathway by which TGF beta regulates sGC expression in PASMC.