Impaired local regulation of ryanodine receptor type 2 by protein phosphatase 1 promotes atrial fibrillation

Cardiovasc Res. 2014 Jul 1;103(1):178-87. doi: 10.1093/cvr/cvu123. Epub 2014 May 8.

Abstract

Aims: Altered Ca(2+) handling in atrial fibrillation (AF) has been associated with dysregulated protein phosphatase 1 (PP1) and subcellular heterogeneities in protein phosphorylation, but the underlying mechanisms remain unclear. This is due to a lack of investigation into the local, rather than global, regulation of PP1 on different subcellular targets such as ryanodine receptor type 2 (RyR2), especially in AF.

Methods and results: We tested the hypothesis that impaired local regulation of PP1 causes RyR2 hyperphosphorylation thereby promoting AF susceptibility. To specifically disrupt PP1's local regulation of RyR2, we used the spinophilin knockout (Sp(-/-)) mice (Mus musculus) since PP1 is targeted to RyR2 via spinophilin. Without spinophilin, the interaction between PP1 and RyR2 was reduced by 64%, while RyR2 phosphorylation was increased by 43% at serine (S)2814 but unchanged at S2808. Lipid bilayer experiments revealed that single RyR2 channels isolated from Sp(-/-) hearts had an increased open probability. Likewise, Ca(2+) spark frequency normalized to sarcoplasmic reticulum Ca(2+) content was also enhanced in Sp(-/-) atrial myocytes, but normalized by Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) inhibitors KN-93 and AIP and also by genetic inhibition of RyR2 S2814 phosphorylation. Finally, Sp(-/-) mice exhibited increased atrial ectopy and susceptibility to pacing-induced AF, both of which were also prevented by the RyR2 S2814A mutation.

Conclusion: PP1 regulates RyR2 locally by counteracting CaMKII phosphorylation of RyR2. Decreased local PP1 regulation of RyR2 contributes to RyR2 hyperactivity and promotes AF susceptibility. This represents a novel mechanism for subcellular modulation of calcium channels and may represent a potential drug target of AF.

Keywords: Atrial fibrillation; Ca2+/calmodulin-dependent kinase II; Protein phosphatase 1; Ryanodine receptor type 2; Spinophilin.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Substitution
  • Animals
  • Atrial Fibrillation / etiology*
  • Atrial Fibrillation / genetics
  • Atrial Fibrillation / metabolism*
  • Calcium Signaling
  • Disease Models, Animal
  • Female
  • Male
  • Mice
  • Mice, Inbred C57BL
  • Mice, Knockout
  • Mice, Mutant Strains
  • Microfilament Proteins / deficiency
  • Microfilament Proteins / genetics
  • Mutant Proteins / genetics
  • Mutant Proteins / metabolism
  • Myocardium / metabolism
  • Nerve Tissue Proteins / deficiency
  • Nerve Tissue Proteins / genetics
  • Phosphorylation
  • Protein Phosphatase 1 / metabolism*
  • Ryanodine Receptor Calcium Release Channel / genetics
  • Ryanodine Receptor Calcium Release Channel / metabolism*
  • Sarcoplasmic Reticulum / metabolism

Substances

  • Microfilament Proteins
  • Mutant Proteins
  • Nerve Tissue Proteins
  • Ryanodine Receptor Calcium Release Channel
  • neurabin
  • ryanodine receptor 2. mouse
  • Protein Phosphatase 1