Β-Adrenergic regulation of cardiac type 2A protein phosphatase through phosphorylation of regulatory subunit B56δ at S573. (February 2018)

• Record Type: Journal Article
• Authors: Ranieri, Antonella; Kemp, Elizabeth; Burgoyne, Joseph R.; Avkiran, Metin
• Is Part Of: Journal of molecular and cellular cardiology. Volume 115(2018)
• Page Start: 20
• Page End: 31
• Publication Date: 2018-02
• Subjects: ARVM adult rat ventricular myocytes -- CaMKII calcium/calmodulin-dependent kinase II -- cMyBP-C cardiac myosin binding protein-C -- cTnI cardiac troponin I -- I-1 inhibitor of protein phosphatase-1 -- ISO isoprenaline -- KO knock out -- MOI multiplicity of infection -- BNZ N6-benzoyl-cAMP -- PLB phospholamban -- PKA protein kinase A -- Ser, S serine -- Thr, T threonine -- PP1 type 1 phosphatase -- PP2A type 2A protein phosphatase -- WT wild type -- βAR β-Adrenergic receptor; β-Adrenergic signaling -- B56δ -- Cardiomyocytes -- Phosphatase -- Phosphorylation -- Type 2A protein phosphatase (PP2A) -- Protein kinase A (PKA); Cardiology -- Periodicals; Heart Diseases -- Periodicals; Molecular Biology -- Periodicals; Cardiologie -- Périodiques; Cardiology; Electronic journals; Periodicals; 616.12
• DOI: 10.1016/j.yjmcc.2017.12.016
• Languages: English
• ISSNs: 0022-2828

Abstract

Abstract: Background: Type 2A protein phosphatase (PP2A) enzymes are serine/threonine phosphatases which comprise a scaffold A subunit, a regulatory B subunit and a catalytic C subunit, and have been implicated in the dephosphorylation of multiple cardiac phosphoproteins. B subunits determine subcellular targeting, substrate specificity and catalytic activity, and can themselves be regulated by post-translational modifications. We explored potential β-adrenergic regulation of PP2A in cardiomyocytes through phosphorylation of the regulatory B subunit isoform B56δ. Methods and results: Phosphate affinity SDS-PAGE and immunoblot analysis revealed increased phosphorylation of B56δ in adult rat ventricular myocytes (ARVM) exposed to the β-adrenergic receptor (βAR) agonist isoprenaline (ISO). Phosphorylation of B56δ occurred at S573, primarily through stimulation of the β1 AR subtype, and was dependent on PKA activity. The functional role of the phosphorylation was explored in ARVM transduced with adenoviruses expressing wild type (WT) or non-phosphorylatable (S573A) B56δ, fused to GFP at the N-terminus. C subunit expression was increased in ARVM expressing GFP-B56δ-WT or GFP-B56δ-S573A, both of which co-immunoprecipitated with endogenous C and A subunits. PP2A activity in cell lysates was increased in response to ISO in ARVM expressing GFP-B56δ-WT but not GFP-B56δ-S573A. Immunoblot analysis of the phosphoproteome in ARVM expressing GFP-B56δ-WT or GFP-B56δ-S573A with antibodies detecting (i) phospho-serine/threonine residues in distinct kinase substrate motifs or (ii) specific phosphorylated residues of functional importance in selected proteins revealed a comparable phosphorylation profile in the absence or presence of ISO stimulation. Conclusions: In cardiomyocytes, βAR stimulation induces PKA-mediated phosphorylation of the PP2A regulatory subunit isoform B56δ at S573, which increases associated PP2A catalytic activity. This is likely to regulate the phosphorylation status of specific B56δ-PP2A substrates, which remain to be identified. Highlights: PP2A subunit B56δ is phosphorylated on β-adrenergic stimulation of cardiomyocytes. Phosphorylation occurs at Ser573 and increases B56δ-PP2A catalytic activity. Response is mediated by the β1 -adrenoceptor subtype and protein kinase A. Phosphorylated B56δ abundance is increased in pathological cardiac hypertrophy.