Abstract 15864: Ring Finger Protein RNF207: A Novel Regulator of HERG Conserved During Vertebrate Evolution
Introduction: The cell surface expression and function of ion channels depend on the balance between biosynthesis, proper folding, forward trafficking, recycling, and degradation. Genome-wide association studies recently reported several novel loci associated with prolongation of the QT interval, one of which is in linkage disequilibrium with the open reading frame of ring finger protein 207 (RNF207), a putative ubiquitin ligase highly expressed in heart. The objective of this study was to investigate the role of RNF207 on action potential formation in vivo and in vitro. Since ubiquitin ligases are often involved in protein turnover, we hypothesized that cardiac RNF207 modulates action potential duration (APD) by affecting one or more underlying ion current(s).
Methods: APD and conduction velocity were assessed by electrophysiological studies of atrial and ventricular myocardium of zebrafish embryos microinjected with morpholinos to knock down expression of endogenous RNF207. Optical mapping of neonatal rabbit cardiomyocytes (NRbCM) adenovirally infected was performed to measure APD. To study the effect of RNF207 on HERG expression, surface biotinylation and Western blot assays were performed using HEK cells. Co-immunoprecipitation and immunofluorescence were applied to study RNF207-binding proteins.
Results: RNF207 zebrafish morphants showed significant prolongation of APD at 48 hours post fertilization (hpf) (mean ventricular APD80 ± S.E.M.: morphants 283±10 ms vs. WT 247±9 ms, p<0.05; atrium: morphants 136±5 ms vs. WT 120±4 ms, p<0.05). In NRbCM, RNF207 significantly shortened APD by approximately 15% compared to control (p<0.05). Overexpression of RNF207 in HEK cells increased surface expression of HERG by 30% (p<0.05). Furthermore an interaction and co-localization of RNF207 with HERG as well as with HSP70, a key chaperone known to increase the expression of HERG, was observed.
Conclusion: RNF207 may contribute to regulate folding/trafficking of HERG, thereby determining action potential duration and cardiac cell excitation.
- © 2013 by American Heart Association, Inc.