IKS, the slowly activating delayed rectifier potassium (K+) current in the heart is critical importance to human physiology as evident from the fact that mutations in either its ? (KCNQ1) or ? (KCNE1) subunit have been linked to multiple cardiac arrhythmia syndromes, including long QT syndrome (LQTS); short QT syndrome; and familial atrial fibrillation. The IKS channel is upregulated during sympathetic stimulation by PKA phosphorylation, which contributes critically to the physiological shortening of cardiac action potentials in response to sympathetic nerve activity. This shortening is necessary to ensure adequate ventricular filling time with accompanying increases in heart rate. It is also during sympathetic stimulation that most sudden deaths from LQTS occur. Understanding the mechanisms that underlie these mutation-induced arrhythmia syndromes requires unraveling the molecular interactions between KCNQ1 and KCNE1 within the context of normal and disease altered IKS channels. But to date, the critical questions of how KCNE1 alters KCNQ1 channel gating and how IKS channels are modulated by PKA are still not fully answered. Previous studies suggest a possible interaction between the N-terminus of KCNQ1 and C-terminus of KCNE1 during adrenergic responses. Here, we will use fluorescent unnatural amino acids as the basis for FRET experiments that will assay the proximity of these critical intracellular domains with and without adrenergic challenge. Our previous work has revealed that ?-AR regulation of channels requires assembly of a macromolecular complex that includes both KCNQ1 and KCNE1, as well as the adaptor protein Yotiao (AKAP 9). We will here use novel nanobodies to deliver regulatory domains of PKA directly to the KCNQ1/KCNE1 channel with and without co-assembly with AKAP9. These experiments will allow dissection of the critical role of AKAP9 in the delivery of signaling molecules to KCNQ1/KCNE1 from additional putative modulatory roles of the AKAP in modulating channel function post phosphorylation. In the recent CryoEM structure of KCNQ1 putative interacting residues between KCNQ1 and KCNE1 map between the VSD and PD, suggesting that KCNE1 is located in this area of the KCNQ1 structure. We will test whether KCNQ1 and KCNQ1/KCNE1 channels open using different gating hinges in S6. We will here also identify KCNQ1-KCNE1 interacting residues and determine whether these residues affect the different gating hinges. PKA has been shown to alter the voltage dependence, sub-conductance occupancy, and kinetics of IKS channels. Using voltage clamp fluorometry together with mutations and PIP2 depletion that uncouple the VSD and PD, we will determine whether PKA affect the VSD, PD, and/or VSD-to-PD coupling in IKS channels. The anticipated results of these experiments will provide a structural basis for control by PKA and KCNE1 of the physiological function of this critical ion channel and will also provide novel targets for the development of drugs to modulate its activity. This would be a milestone toward mutation-specific treatments of diseases, such as cardiac arrhythmias, caused by mutations in KCNQ1 and KCNE1.
Mutations and variants in the IKS potassium channel ? (KCNQ1) and ? (KCNE1) subunits as well as the adaptor protein AKAP9 which coordinates the IKS macromolecular signaling complex have been implicated in a number of life threatening cardiac arrhythmias. Here we will here determine how the IKS channel is regulated by PKA and how assembly with different numbers of its ancillary subunits contributes to normal and pathological function.
|Lassuthova, Petra; Rebelo, Adriana P; Ravenscroft, Gianina et al. (2018) Mutations in ATP1A1 Cause Dominant Charcot-Marie-Tooth Type 2. Am J Hum Genet 102:505-514|
|Bohannon, Briana M; Perez, Marta E; Liin, Sara I et al. (2018) ?-6 and ?-9 polyunsaturated fatty acids with double bonds near the carboxyl head have the highest affinity and largest effects on the cardiac IKs potassium channel. Acta Physiol (Oxf) :e13186|
|Liin, Sara I; Yazdi, Samira; Ramentol, Rosamary et al. (2018) Mechanisms Underlying the Dual Effect of Polyunsaturated Fatty Acid Analogs on Kv7.1. Cell Rep 24:2908-2918|
|Larsson, Johan E; Larsson, H Peter; Liin, Sara I (2018) KCNE1 tunes the sensitivity of KV7.1 to polyunsaturated fatty acids by moving turret residues close to the binding site. Elife 7:|
|Carmona, Emerson M; Larsson, H Peter; Neely, Alan et al. (2018) Gating charge displacement in a monomeric voltage-gated proton (Hv1) channel. Proc Natl Acad Sci U S A 115:9240-9245|
|Barro-Soria, Rene; Liin, Sara I; Larsson, H Peter (2017) Specificity of M-channel activators: binding or effect? J Physiol 595:605-606|
|Barro-Soria, Rene; Ramentol, Rosamary; Liin, Sara I et al. (2017) KCNE1 and KCNE3 modulate KCNQ1 channels by affecting different gating transitions. Proc Natl Acad Sci U S A 114:E7367-E7376|
|Peng, Gary; Barro-Soria, Rene; Sampson, Kevin J et al. (2017) Gating mechanisms underlying deactivation slowing by two KCNQ1 atrial fibrillation mutations. Sci Rep 7:45911|
|Barro-Soria, Rene; Liin, Sara I; Larsson, H Peter (2017) Using fluorescence to understand ? subunit-NaV channel interactions. J Gen Physiol :|
|Qiu, Feng; Chamberlin, Adam; Watkins, Briana M et al. (2016) Molecular mechanism of Zn2+ inhibition of a voltage-gated proton channel. Proc Natl Acad Sci U S A 113:E5962-E5971|
Showing the most recent 10 out of 20 publications