Old dog, new tricks: novel cardiac targets and stress regulation by protein kinase G

Abstract

The second messenger cyclic guanosine 3'5' monophosphate (cGMP) and its downstream effector protein kinase G (PKG) have been discovered more than 40 years ago. In vessels, PKG1 induces smooth muscle relaxation in response to nitric oxide signalling and thus lowers systemic and pulmonary blood pressure. In platelets, PKG1 stimulation by cGMP inhibits activation and aggregation, and in experimental models of heart failure (HF), PKG1 activation by inhibiting cGMP degradation is protective. The net effect of the above-mentioned signalling is cardiovascular protection. Yet, while modulation of cGMP-PKG has entered clinical practice for treating pulmonary hypertension or erectile dysfunction, translation of promising studies in experimental HF to clinical success has failed thus far. With the advent of new technologies, novel mechanisms of PKG regulation, including mechanosensing, redox regulation, protein quality control, and cGMP degradation, have been discovered. These novel, non-canonical roles of PKG1 may help understand why clinical translation has disappointed thus far. Addressing them appears to be a requisite for future, successful translation of experimental studies to the clinical arena.

Keywords: Cardiac mechanosensing; Phosphodiesterase; Proteasome; Protein kinase G; Redox regulation; cGMP-dependent protein kinase type 1.

Figure 1

cGMP-PKG signalling in the cardiomyocyte. cGMP is generated by NO stimulation of soluble guanylate cyclase (sGC) in the cytosol (left) or natriuretic peptide receptor (NPR) coupled particulate guanylate cyclase (pGC) activation at the cell membrane (right). PDE5, which is located at the z-discs, appears to mainly target sGC-synthetized cGMP. PKG attenuates G-protein-coupled signalling (GPCR) via phosphorylation of regulator of G-protein signalling (RGS). Studies suggest that this happens through both sGC- and pGC-activated PKG pools.^, NO-derived cGMP/PKG improves proteasomal function, dampens ischaemia- and doxorubicin-related damaging mitochondrial signalling,^, and reduces activation of the maladaptive transcription factors NFAT and myocyte enhancer factor-2 (MEF2). NO/sGC/cGMP-activated PKG also phosphorylates several myofilament proteins effecting enhanced lusitropy and attenuated beta-adrenergic inotropy.^– PKG stimulated by both cGMP generated via the pGC pathway at the cell membrane or PDE5 inhibition with sildenafil inhibits TRPC6. Calcium that enters the cell through TRPC6 partakes in myofilament activation and force generation (slow force response, SFR), and calcineurin (CaN)-NFAT signalling,^, and possibly mitochondrial calcium and permeability transition. In addition, pGC-cGMP-stimulated PKG selectively enhances transcription factors that have been linked to cell survival and enhanced adaption, GATA-binding protein 4 (GATA4), and cAMP response element-binding protein (CREB). NP-stimulated cGMP is preferentially targeted by PDE9, which localizes close to t-tubules. Oxidation (ROS) attenuates PKG activation and protective signalling in the cardiomyocyte. Whether oxidative activation differs according to the source of cGMP/PKG remains to be determined. PKG prevents activation and nuclear transport of mothers against decapentaplegic homologue 3 (SMAD3) and hence TGFβ-related signalling in myofibroblasts.^, Whether this happens in cardiomyocytes and whether this is differentially impacted according to PKG generation are unclear. Please note that research into the cardiac targets of the PDE9-related cGMP-PKG signal is as yet limited, and many of the studies to date did not strictly differentiate between sGC- and pGC-regulated cGMP signalling. Future studies will reveal the amount of crosstalk between these functional compartments and the respective importance in varied pathophysiologies, and thus paint a more complete or modified picture of compartmentalized PKG-associated signalling.

Figure 2

Oxidative regulation of PKG in smooth and cardiac muscle cells. In smooth muscle cells, oxidant activation of PKG (cysteine 42 disulfide formation) induces PKG translocation to the cell membrane where it phosphorylates Ca²⁺-activated potassium channels (BK_Ca) and induces hyperpolarization and vasorelaxation. Preventing oxidant PKG activation by substituting cysteine 42 with serine (C42S, redox-dead PKG) impairs this process. cGMP activation of PKG is still present; however, it appears that oxidant and cGMP activation impede each other (NTG: nitroglycerine). In contrast, in cardiomyocytes, oxidant activation of PKG, such as happens with neurohumoral activation (Gαq) or pressure overload (TAC) retains oxidized PKG (PKGox) in the cytosol. Redox-dead PKG^C42S, which may still be activated by cGMP, localizes to the cell membrane where it attenuates TRPC6-related calcium entry and associated maladaptive signalling.