Abstract：
Three isoforms of small conductance, calcium-activated potassium (SK) channel subunits have been identified (SK1-3) that exhibit a broad and overlapping tissue distribution. SK channels have been implicated in several disease states including hypertension and atrial fibrillation, but therapeutic targeting of SK channels is hampered by a lack of subtype-selective inhibitors. This is further complicated by studies showing that SK1 and SK2 preferentially form heteromeric channels during co-expression, likely limiting the function of homomeric channels in vivo. Here, we utilized a simplified expression system to investigate functional current produced when human (h) SK2 and hSK3 subunits are co-expressed. When expressed alone, hSK3 subunits were more clearly expressed on the cell surface than hSK2 subunits. hSK3 surface expression was reduced by co-transfection with hSK2. Whole-cell recording showed homomeric hSK3 currents were larger than homomeric hSK2 currents or heteromeric hSK2:hSK3 currents. The smaller amplitude of hSK2:hSK3-mediated current when compared with homomeric hSK3-mediated current suggests hSK2 subunits regulate surface expression of heteromers. Co-expression of hSK2 and hSK3 subunits produced a current that arose from a single population of heteromeric channels as exhibited by an intermediate sensitivity to the inhibitors apamin and UCL1684. Co-expression of the apamin-sensitive hSK2 subunit and a mutant, apamin-insensitive hSK3 subunit [hSK3(H485N)], produced an apamin-sensitive current. Concentration-inhibition relationships were best fit by a monophasic Hill equation, confirming preferential formation of heteromers. These data show that co-expressed hSK2 and hSK3 preferentially form heteromeric channels and suggest that the hSK2 subunit acts as a chaperone, limiting membrane expression of hSK2:hSK3 heteromeric channels.
摘要：
小电导的三个同工型，已经鉴定了钙激活钾（SK）通道亚基（SK1-3），它们表现出广泛而重叠的组织分布。SK通道与几种疾病状态有关，包括高血压和心房颤动，但是缺乏亚型选择性抑制剂阻碍了SK通道的治疗靶向。研究表明SK1和SK2在共表达过程中优先形成异聚通道，可能限制了体内同源通道的功能。这里,我们利用简化的表达系统来研究人(h)SK2和hSK3亚基共表达时产生的功能电流.当单独表达时，hSK3亚基比hSK2亚基在细胞表面更清楚地表达。通过与hSK2共转染降低hSK3表面表达。全细胞记录显示同型hSK3电流大于同型hSK2电流或异型hSK2:hSK3电流。当与同源hSK3介导的电流相比时,hSK2:hSK3介导的电流的较小幅度表明hSK2亚基调节异聚体的表面表达。hSK2和hSK3亚基的共表达产生了由异聚通道的单个群体产生的电流，如对抑制剂apamin和UCL1684的中等敏感性所示。阿帕明敏感hSK2亚基和突变体的共表达，对阿帕明不敏感的hSK3亚基[hSK3（H485N）]产生对阿帕明敏感的电流。浓度-抑制关系最好用单相希尔方程拟合，确认异聚体的优先形成。这些数据表明，共表达的hSK2和hSK3优先形成异聚通道，并表明hSK2亚基充当伴侣，hSK2的限制性膜表达：hSK3异聚通道。
