The deubiquitinase ubiquitin–specific protease 20 is a positive modulator of myocardial β1-adrenergic receptor expression and signaling

Reversible ubiquitination of G protein-coupled receptors regulates their trafficking and signaling; whether deubiquitinases regulate myocardial β₁-adrenergic receptors (β₁ARs) is unknown. We report that ubiquitin-specific protease 20 (USP20) deubiquitinates and attenuates lysosomal trafficking of the β₁AR. β₁AR-induced phosphorylation of USP20 Ser-333 by protein kinase A-α (PKAα) was required for optimal USP20-mediated regulation of β₁AR lysosomal trafficking. Both phosphomimetic (S333D) and phosphorylation-impaired (S333A) USP20 possess intrinsic deubiquitinase activity equivalent to WT activity. However, unlike USP20 WT and S333D, the S333A mutant associated poorly with the β₁AR and failed to deubiquitinate the β₁AR. USP20-KO mice showed normal baseline systolic function but impaired β₁AR-induced contractility and relaxation. Dobutamine stimulation did not increase cAMP in USP20-KO left ventricles (LVs), whereas NKH477-induced adenylyl cyclase activity was equivalent to WT. The USP20 homolog USP33, which shares redundant roles with USP20, had no effect on β₁AR ubiquitination, but USP33 was up-regulated in USP20-KO hearts suggesting compensatory regulation. Myocardial β₁AR expression in USP20-KO was drastically reduced, whereas β₂AR expression was maintained as determined by radioligand binding in LV sarcolemmal membranes. Phospho-USP20 was significantly increased in LVs of wildtype (WT) mice after a 1-week catecholamine infusion and a 2-week chronic pressure overload induced by transverse aortic constriction (TAC). Phospho-USP20 was undetectable in β₁AR KO mice subjected to TAC, suggesting a role for USP20 phosphorylation in cardiac response to pressure overload. We conclude that USP20 regulates β₁AR signaling in vitro and in vivo Additionally, β₁AR-induced USP20 phosphorylation may serve as a feed-forward mechanism to stabilize β₁AR expression and signaling during pathological insults to the myocardium.