Identification of a Protein Phosphatase-1/Phospholamban Complex That Is Regulated by cAMP-Dependent Phosphorylation

In human and experimental heart failure, the activity of the type 1 phosphatase is significantly increased, associated with dephosphorylation of phospholamban, inhibition of the sarco(endo)plasmic reticulum Ca 2+ transport ATPase (SERCA2a) and depressed function. In the current study, we investigated the molecular mechanisms controlling protein phosphatase-1 activity. Using recombinant proteins and complementary in vitro binding studies, we identified a multi-protein complex centered on protein phosphatase-1 that includes its muscle specific glycogen-targeting subunit G M and substrate phospholamban. G M interacts directly with phospholamban and this association is mediated by the cytosolic regions of the proteins. Our findings suggest the involvement of G M in mediating formation of the phosphatase-1/G M /phospholamban complex through the direct and independent interactions of G M with both protein phosphatase-1 and phospholamban. Importantly, the protein phosphatase-1/G M /phospholamban complex dissociates upon protein kinase A phosphorylation, indicating its significance in the β-adrenergic signalling axis. Moreover, protein phosphatase-1 activity is regulated by two binding partners, inhibitor-1 and the small heat shock protein 20, Hsp20. Indeed, human genetic variants of inhibitor-1 (G147D) or Hsp20 (P20L) result in reduced binding and inhibition of protein phosphatase-1, suggesting aberrant enzymatic regulation in human carriers. These findings provide insights into the mechanisms underlying fine-tuned regulation of protein phosphatase-1 and its impact on the SERCA2/phospholamban interactome in cardiac function.