Evidence for protein phosphatase inhibitor-1 playing an amplifier role in beta-adrenergic signaling in cardiac myocytes

The protein phosphatase inhibitor-1 (PPI-1) inhibits phosphatase type-1 (PP1) only when phosphorylated by protein kinase A and could play a pivotal role in the phosphorylation/dephosphorylation balance. Rat cardiac PPI-1 was cloned by reverse transcriptase-polymerase chain reaction, expressed in Eschericia coli, evaluated in phosphatase assays, and used to generate an antiserum. An adenovirus was constructed encoding PPI-1 and green fluorescent protein (GFP) under separate cytomegalovirus promotors (AdPPI-1/GFP). A GFP-only virus (AdGFP) served as control. Engineered heart tissue (EHT) from neonatal rat cardiomyocytes and adult rat cardiac myocytes (ARCMs) were used as model systems. PPI-1 expression was determined in human ventricular samples by Northern blots. Compared with AdGFP, AdPPI-1/GFP-infected neonatal rat cardiomyocytes displayed a 73% reduction in PP1 activity. EHTs infected with AdPPI-1/GFP exhibited a fivefold increase in isoprenaline sensitivity. AdPPI-1/GFP-infected ARCMs displayed enhanced cell shortening as well as enhanced phospholamban phosphorylation when stimulated with 1 nM isoprenaline. PPI-1 mRNA levels were reduced by 57+/-12% in failing hearts with dilated and ischemic cardiomyopathy (n=8 each) compared with nonfailing hearts (n=8). In summary, increased PPI-1 expression enhances myocyte sensitivity to isoprenaline, indicating that PPI-1 acts as an amplifier in beta-adrenergic signaling. Decreased PPI-1 in failing human hearts could participate in desensitization of the cAMP pathway.