A novel human R25C-phospholamban mutation is associated with super-inhibition of calcium cycling and ventricular arrhythmia

Depressed sarcoplasmic reticulum (SR) Ca²⁺ cycling, a universal characteristic of human and experimental heart failure, may be associated with genetic alterations in key Ca²⁺-handling proteins. In this study, we identified a novel PLN mutation (R25C) in dilated cardiomyopathy (DCM) and investigated its functional significance in cardiomyocyte Ca²⁺-handling and contractility.

Exome sequencing identified a C73T substitution in the coding region of PLN in a family with DCM. The four heterozygous family members had implantable cardiac defibrillators, and three developed prominent ventricular arrhythmias. Overexpression of R25C-PLN in adult rat cardiomyocytes significantly suppressed the Ca²⁺ affinity of SR Ca²⁺-ATPase (SERCA2a), resulting in decreased SR Ca²⁺ content, Ca²⁺ transients, and impaired contractile function, compared with WT-PLN. These inhibitory effects were associated with enhanced interaction of R25C-PLN with SERCA2, which was prevented by PKA phosphorylation. Accordingly, isoproterenol stimulation relieved the depressive effects of R25C-PLN in cardiomyocytes. However, R25C-PLN also elicited increases in the frequency of Ca²⁺ sparks and waves as well as stress-induced aftercontractions. This was accompanied by increased Ca²⁺/calmodulin-dependent protein kinase II activity and hyper-phosphorylation of RyR2 at serine 2814.

The findings demonstrate that human R25C-PLN is associated with super-inhibition of SERCA2a and Ca²⁺ transport as well as increased SR Ca²⁺ leak, promoting arrhythmogenesis under stress conditions. This is the first mechanistic evidence that increased PLN inhibition may impact both SR Ca²⁺ uptake and Ca²⁺ release activities and suggests that the human R25C-PLN may be a prognostic factor for increased ventricular arrhythmia risk in DCM carriers.