Dataset Information

Individual IKs channels at the surface of mammalian cells contain two KCNE1 accessory subunits.

ABSTRACT: KCNE1 (E1) ?-subunits assemble with KCNQ1 (Q1) voltage-gated K(+) channel ?-subunits to form IKslow (IKs) channels in the heart and ear. The number of E1 subunits in IKs channels has been an issue of ongoing debate. Here, we use single-molecule spectroscopy to demonstrate that surface IKs channels with human subunits contain two E1 and four Q1 subunits. This stoichiometry does not vary. Thus, IKs channels in cells with elevated levels of E1 carry no more than two E1 subunits. Cells with low levels of E1 produce IKs channels with two E1 subunits and Q1 channels with no E1 subunits--channels with one E1 do not appear to form or are restricted from surface expression. The plethora of models of cardiac function, transgenic animals, and drug screens based on variable E1 stoichiometry do not reflect physiology.

SUBMITTER: Plant LD

PROVIDER: S-EPMC3986162 | biostudies-other | 2014 Apr

REPOSITORIES: biostudies-other

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Publications

Individual IKs channels at the surface of mammalian cells contain two KCNE1 accessory subunits.

Plant Leigh D LD Xiong Dazhi D Dai Hui H Goldstein Steve A N SA

Proceedings of the National Academy of Sciences of the United States of America 20140303 14

KCNE1 (E1) β-subunits assemble with KCNQ1 (Q1) voltage-gated K(+) channel α-subunits to form IKslow (IKs) channels in the heart and ear. The number of E1 subunits in IKs channels has been an issue of ongoing debate. Here, we use single-molecule spectroscopy to demonstrate that surface IKs channels with human subunits contain two E1 and four Q1 subunits. This stoichiometry does not vary. Thus, IKs channels in cells with elevated levels of E1 carry no more than two E1 subunits. Cells with low leve ...[more]

PMID: 24591645

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Project description:Arachidonic acid (AA) inhibits the activity of several different voltage-gated Ca(2+) channels by an unknown mechanism at an unknown site. The Ca(2+) channel pore-forming subunit (Ca(V)alpha(1)) is a candidate for the site of AA inhibition because T-type Ca(2+) channels, which do not require accessory subunits for expression, are inhibited by AA. Here, we report the unanticipated role of accessory Ca(V)beta subunits on the inhibition of Ca(V)1.3b L-type (L-) current by AA. Whole cell Ba(2+) currents were measured from recombinant channels expressed in human embryonic kidney 293 cells at a test potential of -10 mV from a holding potential of -90 mV. A one-minute exposure to 10 microM AA inhibited currents with beta(1b), beta(3), or beta(4) 58, 51, or 44%, respectively, but with beta(2a) only 31%. At a more depolarized holding potential of -60 mV, currents were inhibited to a lesser degree. These data are best explained by a simple model where AA stabilizes Ca(V)1.3b in a deep closed-channel conformation, resulting in current inhibition. Consistent with this hypothesis, inhibition by AA occurred in the absence of test pulses, indicating that channels do not need to open to become inhibited. AA had no effect on the voltage dependence of holding potential-dependent inactivation or on recovery from inactivation regardless of Ca(V)beta subunit. Unexpectedly, kinetic analysis revealed evidence for two populations of L-channels that exhibit willing and reluctant gating previously described for Ca(V)2 channels. AA preferentially inhibited reluctant gating channels, revealing the accelerated kinetics of willing channels. Additionally, we discovered that the palmitoyl groups of beta(2a) interfere with inhibition by AA. Our novel findings that the Ca(V)beta subunit alters kinetic changes and magnitude of inhibition by AA suggest that Ca(V)beta expression may regulate how AA modulates Ca(2+)-dependent processes that rely on L-channels, such as gene expression, enzyme activation, secretion, and membrane excitability.

Dataset Information

Individual IKs channels at the surface of mammalian cells contain two KCNE1 accessory subunits.

Publications

Individual IKs channels at the surface of mammalian cells contain two KCNE1 accessory subunits.

Similar Datasets

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