Extended Data Fig. 1: Putative PKA phosphorylation sites in α_1C and β_2B subunits.

a, Left column, the 35 putative PKA phosphorylation sites in rabbit α_1C. Centre, the 51 residues in red are either predicted phosphorylation sites or within the immediate region of the predicted phosphorylation sites. Right, all 51 residues were replaced with alanine in 35-mutant α_1C transgenic mice. b, Combined bar and column scatter plot of Boltzmann function parameters, V₅₀. Data are mean ± s.e.m. **P < 0.01; ***P < 0.001; ****P < 0.0001 by paired two-tailed t-test. pWT α, n = 19; 35-α mutant, n = 14; 28-β mutant, n = 16; 35-α mutant × 28-β mutant, n = 24. Specific P values can be found in the associated Source Data (see Supplementary Information). c, Graph showing isoproterenol- and forskolin-induced increases in nisoldipine-resistant current, stratified by total basal current density before nisoldipine treatment. d, Left, the 28 putative PKA phosphorylation sites in the N-terminal (NT), Hook, GK and C-terminal (CT) domains of β_2B. Centre, the 37 residues in red are either predicted phosphorylation sites or within the immediate vicinity of predicted phosphorylation sites, and were mutated to alanine in the 28-mutant GFP-tagged β_2B transgenic mice (right). e, Fluorescence imaging of isolated cardiomyocytes expressing the GFP-tagged 28-β mutant. Representative of images from more than five biologically independent mice. f, Anti-β-subunit immunoblot of cleared lysates from doxycycline-fed 35-mutant α_1C transgenic (TG) mice or 35-mutant α_1C × GFP-tagged 28-mutant β_2B expressing mice hearts. Representative of immunoblots obtained from at least three biologically independent mice. g, Anti-Flag antibody (upper) and anti-β antibody (lower) immunoblots of anti-Flag antibody immunoprecipitations from cleared lysates of hearts from pWT, 35-α and three mice expressing 35-α × GFP-tagged-28-β. Representative images from two independent experiments. For source gel data, see Supplementary Fig. 1.

Source data