Pressure-Overload–Induced Subcellular Relocalization/Oxidation of Soluble Guanylyl Cyclase in the Heart Modulates Enzyme Stimulation

Abstract

Rationale Soluble guanylyl cyclase (sGC) generates cyclic guanosine monophophate (cGMP) upon activation by nitric oxide (NO). Cardiac NO-sGC-cGMP signaling blunts cardiac stress responses, including pressure-overload induced hypertrophy. The latter itself depresses signaling via this pathway by reducing NO generation and enhancing cGMP hydrolysis. Objective We tested the hypothesis that the sGC response to NO also declines with pressure-overload stress, and assessed the role of heme-oxidation and altered intracellular compartmentation of sGC as potential mechanisms. Methods and Results C57BL/6 mice subjected to transverse aortic constriction developed cardiac hypertrophy and dysfunction. NO-stimulated sGC activity was markedly depressed, while NO- and heme-independent sGC activation by BAY 60-2770 was preserved. Total sGCα1 and β1 expression were unchanged by TAC, however sGCβ1 subunits shifted out of caveolin-enriched microdomains. NO-stimulated sGC activity was 2–3-fold greater in Cav3-containing lipid raft versus non-lipid raft domains in control, and 6-fold greater after TAC. In contrast, BAY 60-2770 responses were >10 fold higher in non-Cav3 domains with and without TAC, declining about 60% after TAC within each compartment. Mice genetically lacking Cav3 had reduced NO and BAY-stimulated sGC activity in microdomains containing Cav3 for controls, but no change within non-Cav3-enriched domains. Conclusions Pressure-overload depresses NO/heme-dependent sGC activation in the heart, consistent with enhanced oxidation. The data reveal a novel additional mechanism for reduced NO-coupled sGC activity related to dynamic shifts in membrane microdomain localization, with Cav3-microdomains protecting sGC from heme-oxidation and facilitating NO-responsiveness. Translocation of sGC out of this domain favors sGC oxidation and contributes to depressed NO-stimulated sGC activity.

Transgenic Mice for Real-Time Visualization of cGMP in Intact Adult Cardiomyocytes

“…localization of the sensor. In contrast, previously published data documented small but significant increases of cardiomyocyte cGMP after NO donor stimulation in C57BL/6 myocardium, 40 which was also the case in our working heart experiments. We also tested the possibility that the isolation procedure might lead to oxidation and loss of NO sensitivity of NO-GC.…”

Preconditioning with soluble guanylate cyclase activation prevents postischemic inflammation and reduces nitrate tolerance in heme oxygenase-1 knockout mice

“…Moreover, the tethering of sGC near a source of antioxidant activity would convey protection from Fe 2ϩ oxidation. Such protection was compromised in pressureoverloaded hearts, which exhibited reduced NO-stimulated sGC activity (41). The reduction was associated with translocation of sGC from caveolin-3 to cytosolic fractions where oxidation of sGC occurred more readily.…”

Prevention of PKG1α oxidation augments cardioprotection in the stressed heart

“…Despite enthusiasm for the therapeutic potential of PKG enhancement strategies, their impact has been somewhat variable, raising concerns that cGMP/PKG signaling may become blunted by disease modifiers such as oxidative stress (7), which depresses NO (7) and cGMP synthesis (8,9). Studies have also shown that oxidative stress impacts PKG1α, the primary cardiovascular isoform.…”