Abstract 657: A Tetramerization Domain Mutation in KCNA5 Identified in Pulmonary Arterial Hypertension Patients Causes Dysfunctional K+Currents
Pulmonary vasoconstriction and pulmonary vascular remodeling are major causes for the elevated pulmonary vascular resistance in patients with idiopathic pulmonary arterial hypertension (IPAH). Activity of voltage-gated K+(Kv) channels regulates the resting membrane potential (Em) in pulmonary artery smooth muscle cells (PASMC), and is involved in the regulation of PASMC contraction, proliferation and apoptosis. The highly conserved T1 domain in the N-terminus of Kv channels is important for channel tetramerization, association with regulatory βsubunits, and localization to the plasma membrane. Downregulated expression of Kv channels (e.g., Kv1.5) and decreased Kv current (IK(V)) have been observed in IPAH PASMC compared to normal PASMC. We recently found two non-synonymous mutations, G182R and E211D, in the T1 domain of IPAH patients’ Kv1.5 channel, encoded by the KCNA5 gene. HEK-293 cells transfected with mutant (G182R) or wildtype (WT) Kv1.5 channel revealed similar current amplitudes and activation thresholds (−40 mV). However, kinetic differences between the two channels were found to exist. While WT currents exhibited both fast- and non-inactivating components, the fast inactivating component was absent in G182R-transfected cells. The time to reach maximum current activation when cells were depolarized from −70 to + 60 mV was significantly longer for the G182R currents than for the WT currents (27 vs. 9 ms). Differences in tail currents were also observed: WT tail currents peaked and then slowly inactivated. However, the G182R tail currents inactivated rapidly and were at near resting levels by the end of the repolarizing pulse. Lastly, the two channels’ pharmacological responses to 4-AP, a Kv channel blocker, differed. While WT currents were ~66% inhibited by 5 mM 4-AP, G182R currents were about 90% inhibited and showed a much slower recovery after washout of 4-AP compared to WT. Overall, the G182R mutant channel showed slow activation, fast inactivation, and fast channel closure; these effects would reduce channel activity and cause membrane depolarization. These results indicate that the G182R mutation may be one of the genetic causes for the inhibited Kv channel activity in PASMC from some IPAH patients.