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

Abstract

AimsAltered 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 resultsWe 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 2/2 ) 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 2/2 hearts had an increased open probability.Likewise, Ca 2+ spark frequency normalized to sarcoplasmic reticulum Ca 2+ content was also enhanced in Sp 2/2 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 2/2 mice exhibited increased atrial ectopy and susceptibility to pacing-induced AF, both of which were also prevented by the RyR2 S2814A mutation. ConclusionPP1 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.- ------------------------------ KeywordsAtrial fibrillation † Protein phosphatase 1 † Ryanodine receptor type 2 † Spinophilin † Ca 2+ /calmodulindependent kinase II

The Cardiac Ryanodine Receptor Phosphorylation Hotspot Embraces PKA in a Phosphorylation-Dependent Manner

“…It remains to be tested which exact RyR2 residues can form these interactions and whether phosphorylation of other residues, such as S2807, can also induce a helix. The conformational change may explain the strong functional effect of CaMKII on the activity of RyR2 (Ai et al, 2005;Ather et al, 2013;Chiang et al, 2014;Curran et al, 2007;Di Carlo et al, 2014;Dixit et al, 2013;Kushnir et al, 2010;Respress et al, 2012;Uchinoumi et al, 2016;Wehrens et al, 2004) and to our knowledge describes a unique structural crosstalk between PKA and CaMKII on RyR2.…”

“…Aberrant phosphorylation of RyR2 has been linked to atrial fibrillation (Chiang et al, 2014;Si et al, 2018), heart failure (Dobrev and Wehrens, 2014;Marx et al, 2000;van Oort et al, 2010), increased cell death in ischemia-reperfusion (Di Carlo et al, 2014), cognitive dysfunction (Liu et al, 2012), and Alzheimer's disease (Lacampagne et al, 2017), although the exact molecular mechanisms are heavily disputed (Bers, 2012;Eschenhagen, 2010).…”