Alterations in the Interactome of Serine/Threonine Protein Phosphatase Type-1 in Atrial Fibrillation Patients

ABSTRACT: Objectives: To test the hypothesis that serine/threonine protein phosphatase type-1 (PP1) is dysregulated in paroxysmal atrial fibrillation (pAF) at the level of its regulatory subunits (R-subunits). Background: AF is the most common sustained cardiac arrhythmia yet current pharmacologic treatment is ineffective. PP1, a major phosphatase in the heart, consists of a catalytic subunit (PP1c) and a large set of R-subunits that confer localization and substrate specificity to the holoenzyme. Previous studies suggest that PP1 is dysregulated in AF but the mechanisms are unknown. Methods: Cardiac lysates were co-immunoprecipitated with anti-PP1c antibody followed by mass spectrometry-based (quantitative) profiling of associated R-subunits. Subsequently, label-free quantification was used to evaluate altered R-subunit-PP1c interactions in pAF patients. R-subunits with altered binding to PP1c in pAF were further validated using qRT-PCR, Western blotting (WB), immunocytochemistry, and co-immunoprecipitation. Results: 135 and 78 putative PP1c-interactors were captured respectively from mouse ventricles and human atria, with many previously unreported interactors with conserved PP1c-docking motifs. Increases in binding were found between PP1c and PPP1R7, CSDA, and PDE5A in pAF patients, with CSDA and PDE5A being novel interactors validated by bioinformatics, immunocytochemistry and co-immunoprecipitation. WB confirmed that these upregulated associations cannot be ascribed to changes in global protein expression alone. Conclusion: Subcellular heterogeneity in PP1 activity and downstream protein phosphorylation in AF may be attributed to alterations in PP1c-R-subunits interactions, which impair PP1 targeting to proteins involved in electrical and Ca2+-remodeling. This represents a novel concept in AF pathogenesis and provides highly-specific drug targets for treating AF.

INSTRUMENT(S): LTQ Orbitrap Elite, Q Exactive

ORGANISM(S): Homo Sapiens (human) Mus Musculus (mouse)

TISSUE(S): Heart

DISEASE(S): Heart Conduction Disease

SUBMITTER: Nicolas Lebesgue  

LAB HEAD: Arjen Scholten

PROVIDER: PXD000697 | Pride | 2015-03-30

REPOSITORIES: Pride