PP2A-B56α is a key determinant of cardiac protein phosphorylation and functional responses to β-adrenergic signalling

B56α is a protein phosphatase 2 A (PP2A) regulatory subunit which modulates the heart's inotropic response to acute β-adrenergic receptor (β-AR) stimulation, although knowledge of the underlying molecular mechanisms is limited. In this study, mice deficient for B56α and wildtype controls received an intraperitoneal injection of isoproterenol (0.1 mg/kg) to activate β-AR signalling <i>in vivo</i>, and their hearts examined two minutes post-injection by quantitative phosphoproteomics to identify mechanisms of acute β-adrenergic signalling. We identified site- and genotype-specific phosphorylation changes on >200 proteins, including 25 hyperphosphorylated proteins harbouring a B56 binding motif as putative substrates. Functional enrichment analysis pointed to cardiac Ca²⁺ release and contractility as key processes impacted by B56α deficiency, as well as cardiac muscle hypertrophy as a potential disease mechanism. In vitro, loss of B56α in cardiomyocytes blunted acute isoproterenol-induced increases in intracellular calcium transient amplitude, confirming that B56α plays a key role in calcium handling. In vivo, loss of B56α protected mice from developing systolic dysfunction in response to sustained isoproterenol infusion (60 mg/kg/day for 14 days), despite comparable increases in heart mass. These findings reaffirm a key role for B56α as a mediator of physiologically important cardiac responses to β-AR stimulation and reveal potential new molecular mechanisms for this regulatory function, including putative cardiac B56α substrates.