Panaghie Gianina, Tai Kwok-Keung, Abbott Geoffrey W
Greenberg Division of Cardiology, Department of Medicine, Weill Medical College of Cornell University, 520 East 70th Street, New York, NY 10021, USA.
J Physiol. 2006 Feb 1;570(Pt 3):455-67. doi: 10.1113/jphysiol.2005.100644. Epub 2005 Nov 24.
KCNQ1 alpha subunits form functionally distinct potassium channels by coassembling with KCNE ancillary subunits MinK and MiRP2. MinK-KCNQ1 channels generate the slowly activating, voltage-dependent cardiac IKs current. MiRP2-KCNQ1 channels form a constitutively active current in the colon. The structural basis for these contrasting channel properties, and the mechanisms of alpha subunit modulation by KCNE subunits, are not fully understood. Here, scanning mutagenesis located a tryptophan-tolerant region at positions 338-340 within the KCNQ1 pore-lining S6 domain, suggesting an exposed region possibly amenable to interaction with transmembrane ancillary subunits. This hypothesis was tested using concomitant mutagenesis in KCNQ1 and in the membrane-localized 'activation triplet' regions of MinK and MiRP2 to identify pairs of residues that interact to control KCNQ1 activation. Three pairs of mutations exerted dramatic effects, ablating channel function or either removing or restoring control of KCNQ1 activation. The results place KCNE subunits close to the KCNQ1 pore, indicating interaction of MiRP2-72 with KCNQ1-338; and MinK-59,58 with KCNQ1-339, 340. These data are consistent either with perturbation of the S6 domain by MinK or MiRP2, dissimilar positioning of MinK and MiRP2 within the channel complex, or both. Further, the results suggest specifically that two of the interactions, MiRP2-72/KCNQ1-338 and MinK-58/KCNQ1-340, are required for the contrasting gating effects of MinK and MiRP2.
KCNQ1α亚基通过与KCNE辅助亚基MinK和MiRP2共同组装形成功能不同的钾通道。MinK-KCNQ1通道产生缓慢激活、电压依赖性的心脏IKs电流。MiRP2-KCNQ1通道在结肠中形成组成性激活电流。这些截然不同的通道特性的结构基础以及KCNE亚基对α亚基的调节机制尚未完全了解。在这里，扫描诱变在KCNQ1孔衬S6结构域内的338-340位定位了一个色氨酸耐受区域，表明该暴露区域可能易于与跨膜辅助亚基相互作用。使用KCNQ1以及MinK和MiRP2的膜定位“激活三联体”区域中的伴随诱变来测试该假设，以识别相互作用以控制KCNQ1激活的残基对。三对突变产生了显著影响，消除了通道功能或消除或恢复了对KCNQ1激活的控制。结果表明KCNE亚基靠近KCNQ1孔，表明MiRP2-7与KCNQ1-338相互作用；以及MinK-59、58与KCNQ1-339、340相互作用。这些数据与MinK或MiRP2对S6结构域的扰动、MinK和MiRP2在通道复合物中的不同定位或两者一致。此外，结果特别表明，MinK和MiRP2的对比门控效应需要MiRP2-72/KCNQ1-338和MinK-58/KCNQ1-340这两种相互作用。
