Sulfhydryl-dependent dimerization of soluble guanylyl cyclase modulates the relaxation of porcine pulmonary arteries to nitric oxide

Abstract

The dimeric status of nitric oxide (NO)-sensitive soluble guanylyl cyclase (sGC) is obligatory for its catalyzing activity to synthesis the second messenger cyclic guanosine monophosphate (cGMP), which leads to vasodilatation. The present study was conducted to determine whether or not the dimerization of sGC is modulated by thiol-reducing agents and its influences on relaxation of pulmonary arteries caused by NO. The dimers and monomers of sGC and cGMP-dependent protein kinase (PKG) were analyzed by Western blotting. The intracellular cGMP content was measured by enzyme-linked immunosorbent assay. Relaxations of isolated porcine pulmonary arteries were determined by organ chamber technique. Protein levels of sGC dimers were decreased by thiol reductants dithiothreitol (DTT), reduced L-glutathione, L-cysteine, and tris(2-carboxyethyl)phosphine (TCEP), associated with decreased cGMP elevation, attenuated relaxations to NO. DTT at concentrations that affected sGC dimerization and activity showed no effect on PKG dimerization nor relaxation to 8-Br-cGMP. Hypoxia decreased the dimerization and activity of sGC of the arteries. The suppression of DTT and TCEP on sGC dimerization and activity was augmented by hypoxia. In the presence of DTT and TCEP, relaxations of porcine pulmonary artery caused by NO were significantly less under hypoxia compared with those under normoxia. These results suggest that the dimerization and activity of sGC along with NO-induced vasodilatation can be modulated in a thiol-dependent manner. Such a mechanism may be involved in the diminished response of pulmonary arteries to NO under hypoxia.