E3 ubiquitin ligase rififylin has yin and yang effects on rabbit cardiac transient outward potassium currents (Ito) and corresponding channel proteins

Abstract

Genome-wide association studies (GWAS) have reported a correlation between a single nucleotide polymorphism of the RING finger E3 ubiquitin protein ligase rififylin (RFFL) and QT interval variability in humans (Newton-Cheh et al., 2009). Previously, we have shown that RFFL down-regulates expression and function of the human-like ether-a-go-go-related gene (hERG) potassium channel and corresponding rapidly activating delayed rectifier potassium current (IKr) in adult rabbit ventricular cardiomyocytes (ARbCMs). Here we report that RFFL also affects the transient outward current (Ito), but in a peculiar way. RFFL overexpression in ARbCMs significantly decreases the contribution of its fast component (Ito,f) from 35% to 21%, and increases the contribution of its slow component (Ito,s) from 65% to 79%. Since Ito,f in rabbits is mainly conducted by Kv4.3, we investigated the effect of RFFL on Kv4.3 expressed in HEK293A cells. We found that RFFL overexpression reduced Kv4.3 expression and corresponding Ito,f in a RING domain-dependent manner in the presence or absence of its accessory subunit KChIP2. On the other hand, RFFL overexpression in Kv1.4-expressing HEK cells leads to an increase in both Kv1.4 expression level and Ito,s, similarly in a RING domain-dependent manner. Our physiologically detailed rabbit ventricular myocyte computational model shows that these yin and yang effects of RFFL overexpression on Ito,f, and Ito,s affect phase 1 of the action potential waveform and slightly decrease its duration in addition to suppressing IKr. Thus, RFFL modifies cardiac repolarization reserve via ubiquitination of multiple proteins that differently affect various potassium channels and cardiac action potential duration (APD).