Studies On Cause Of LQTS By KCNQ1 Mutant And Ethanol Effect On KCNQ1 Channel

Ion channel, a fusion protein regulating transmembrane ion flux, is the basis of bio-electrical activity and the membrane excitablity of nerve, muscle and other tissu. Potassium channels are the most various ion channels, and widely exist in skeletal muscle, nerve, heart, vessels, trachea, gastrointestinal tract, blood and endocrine gland cells. KCNQ1 channel, which is an important branch of voltage-gated potassium channels, has a large distribution in the heart, as well as the inner ear. Combined with its auxiliary subunit minK, KCNQ1/minK complex can generate a delayed outward rectifier K⁺ current, Iks. to end the action potential in myocardial cells, therefore, KCNQ1 mutations would cause a dysfunction of the channel, thus give rise to the long QT syndrome (LQTS), eventually lead to serious arrhythmias, ventricular fibrillation and cardiac shock. Mutations in KCNQ gene can lead to many diseases, thus, it is very contributing to clarify the characteristics of KCNQ1 channels, both for the ion channel research and clinical medicine.We studied the mechanism of LQT1 arising from one KCNQ1 mutation, L191P, in HEK293 cells. Over 100 KCNQ1 mutantions can lead to the type-1 LQTS (LQT1). It is reported that KCNQ1-Leu191, which locates in the intracellular S2-S3 linker of KCNQ1, can lead to LQT1 directly, and～16% LQTS-related mutations mustered in this region. By electrophysiological method, we found the currents of L191P/minK channel were much smaller than those of KCNQ1/minK channels. Although the average currents of L191P/minK were reduced to more than half compared with those of WT/minK, but there was no shift in the G-V curves. By immunofluorescence method, we also found L191P channels causing a trafficking deficiency, leading to the minishing of I_Ks and then, LQTS.We found the surface expression decreased with decreasing hydrophobicity of the middle residue 'X' of the RXR motif. Establishing a model of E(?)M, we generalize the Boltzmann formula to explain the results, and we got the essence of LQT1 phenotype through the binomial distribution model, that is the dominant-negative effect.We also studied the blocking mechanism of ethanol (alcohol) on KCNQ1 in Xenopus laevis oocytes. Ethanol has a wide range of pharmacological effects on the human body, but researchers do not understand the mechanism it works. Using two-electrode voltage clamp, we showed that ethanol blocked specificly I_Ks, even the straight-chain n-alcohol also blocked the channel. The chain length was longer, the same concentration blocked stronger. The voltage-dependent and bio-states blocking showed us that ethanol blocked the channel both from outside and inside. Through mutation scanning, we also found that amino acid Ile257 of KCNQ1 played an important role in the blocking.We also used the Medlab bio-signal acquisition and processing system to measure the ECG of the injected mice, and results showed that a certain concentration of ethanol blocked cardiac potassium channels, which delayed action potentials repolarization and thus slowed down the heart rate; and a particular concentration of ethanol (higher over security levels) stimulated the cardiac potassium ion channels opening, thus speeded up the action potential repolarization, thus speeded up the heart rate. We used a "pocket" model to further clarify the interaction beteween n-ethanol and KCNQ1 channel, which will make a graet sense to the study of the pathogenic mechanisms related to heart disease.