Abstract 010: Phosphodiesterase 3a Catalytic-domain Mutants Cause Hypertension With Brachydactyly

Abstract

Mendelian syndromes give great insight into pathogenesis and have implicated salt handling. Hypertension with brachydactyly (HTNB) is unique in that a direct increase in peripheral vascular resistance is produced by activating mutations in phosphodiesterase 3A (PDE3A). A 50 mm Hg blood-pressure elevation by age 50 years causes stroke in untreated persons. Here, we report mutations in the PDE3 catalytic domain found in two new HTNB families. Since structural predictions indicated increased PDE3A cleavage activity, we used Förster resonance energy transfer (FRET) to measure the PDE3-specific cAMP degradation in cytosolic fractions from transfected cells. The newly discovered catalytic-domain PDE3A mutants, R862C and L910P, both showed a shorter transient emission-ratio change upon cAMP addition than PDE3A wildtype, indicating faster cAMP turnover. The V max of all experiments were extracted from FRET data and quantitated and analyzed statistically. The mean values V max (nM (cAMP)/s) were 143±7 for L910P, 63±3 for R862C and 73±4 for T445N, and 45±3 for WT respectively, (p<0.0001, <0.001, <0.05). We found that all disease-relevant mutations led to a clearly hyperactive PDE3A. The L910P catalytic-domain mutation resulted in the most dramatic increase in cAMP hydrolysis. That mutation also exhibited the most stepwise resistance to milrinone. Recently, we showed that cAMP signaling is organized in nanometer-sized cellular compartments. We suggest that the gain-of-function PDE3 mutations result in unphysiologically larger cAMP nanodomains and thereby dysregulate crucially important cAMP nanoarchitecture. Spatiotemporal cAMP signaling and precise PDE-controlled effector activation, could usher in a site-specific nanomolecular pharmacology.