cGMP/PKG signaling pathway has been implicated in regulation of such physiological processes as smooth muscle relaxation and neuronal plasticity. The amplitude and duration of the cGMP signal is controlled by cGMP-specific, cGMP-binding phosphodiesterase (PDE5), which is highly expressed in all types of smooth muscle and cerebellar Purkinje cells. PDE5 has two highly homologous domains in its N-terminal end, recently named GAF A and GAF B based on their sequence homology with similar motifs in a wide group of proteins. The functional roles for these domains in regulation of PDE5 activity has not been fully understood.

Direct PDE5 activation upon cGMP binding to the regulatory GAF domain has been shown in a study of recombinant mouse PDE5 under conditions when the cGMP binding sites were partially or fully occupied. The effect of cGMP occupancy of the cGMP-binding sites on the PDE5 catalytic domain was disrupted by application of mouse monoclonal antibodies, generated against the GAF domain of PDE5 and capable of specific blocking of cGMP binding to the GAF A domain. Pretreatment with this antibody completely prevented PDE5 activation and lowered basal/non-stimulated PDE5 activity revealing that without the stimulatory effect of cGMP binding PDE5 had low intrinsic catalytic activity.

cGMP induced PDE5 activation appeared to be necessary and sufficient to achieve full activation of PDE5 even in the absence of phosphorylation. PKG could phosphorylate PDE5 effectively only in the activated state. The phosphorylation status of PDE5, detected in intact tissues by using phospho-specific PDE5 antibody, correlated with changes in PKG activation. KT5823, a compound considered to be a specific PKG inhibitor, was not able to inhibit PKG induced PDE5 phosphorylation either in smooth muscle cells nor in Purkinje cells, while the role of PKG in PDE5 phosphorylation in these tissues has been proven by using PKG I knockout mice.

Conclusion

Acknowledgements