The LQT-associated calmodulin mutant E141G induces disturbed Ca2+-dependent binding and a flickering gating mode of the CaV1.2 channel.
Calmodulin (CaM) mutations are associated with congenital long QT (LQT) syndrome (LQTS), which may be related to the dysregulation of the cardiac-predominant Ca2+ channel isoform CaV1.2. Among various mutants, CaM-E141G was identified as a critical missense variant. However, the interaction of this CaM mutant with the CaV1.2 channel has not been determined. In this study, by utilizing a semiquantitative pull-down assay, we explored the interaction of CaM-E141G with CaM-binding peptide fragments of the CaV1.2 channel. Using the patch-clamp technique, we also investigated the electrophysiological effects of the mutant on CaV1.2 channel activity. We found that the maximum binding (Bmax) of CaM-E141G to the proximal COOH-terminal region, PreIQ-IQ, PreIQ, IQ, and NT (an NH2-terminal peptide) was decreased (by 17.71-59.26%) compared with that of wild-type CaM (CaM-WT). In particular, the Ca2+-dependent increase in Bmax became slower with the combination of CaM-E141G + PreIQ and IQ but faster in the case of NT. Functionally, CaM-WT and CaM-E141G at 500 nM Ca2+ decreased CaV1.2 channel activity to 24.88% and 55.99%, respectively, compared with 100 nM Ca2+, showing that the inhibitory effect was attenuated in CaM-E141G. The mean open time of the CaV1.2 channel was increased, and the number of blank traces with no channel opening was significantly decreased. Overall, CaM-E141G exhibits disrupted binding with the CaV1.2 channel and induces a flickering gating mode, which may result in the dysfunction of the CaV1.2 channel and, thus, the development of LQTS. The present study is the first to investigate the detailed binding properties and single-channel gating mode induced by the interaction of CaM-E141G with the CaV1.2 channel.
['Su|Jingyang|J|', 'Gao|Qinghua|Q|', 'Yu|Lifeng|L|', 'Sun|Xuanxuan|X|', 'Feng|Rui|R|', 'Shao|Dongxue|D|', 'Yuan|Yuan|Y|', 'Zhu|Zhengnan|Z|', 'Sun|Xuefei|X|', 'Kameyama|Masaki|M|', 'Hao|Liying|L|']
D000818:Animals D002118:Calcium D020746:Calcium Channels, L-Type D020013:Calcium Signaling D002147:Calmodulin D006168:Guinea Pigs D006801:Humans D015640:Ion Channel Gating D007700:Kinetics D008133:Long QT Syndrome D020125:Mutation, Missense D018408:Patch-Clamp Techniques D010455:Peptides D011485:Protein Binding D020033:Protein Isoforms
calmodulin mutant CaV1.2 channel Ca2+-dependent inactivation long QT syndrome