Down-regulation of endogenous KLHL1 decreases voltage-gated calcium current density

Paula P. Perissinotti; Elizabeth G. Ethington; Leanne Cribbs; Michael D. Koob; Jody Martin; Erika S. Piedras-Rentería

doi:10.1016/j.ceca.2014.03.002

Down-regulation of endogenous KLHL1 decreases voltage-gated calcium current density

Paula P. Perissinotti, Elizabeth G. Ethington, Leanne Cribbs, Michael D. Koob, Jody Martin, Erika S. Piedras-Rentería

Laboratory Medicine and Pathology

Research output: Contribution to journal › Article › peer-review

16 Scopus citations

Abstract

The actin-binding protein Kelch-like 1 (KLHL1) can modulate voltage-gated calcium channels in vitro. KLHL1 interacts with actin and with the pore-forming subunits of Cav2.1 and CaV3.2 calcium channels, resulting in up-regulation of P/Q and T-type current density. Here we tested whether endogenous KLHL1 modulates voltage gated calcium currents in cultured hippocampal neurons by down-regulating the expression of KLHL1 via adenoviral delivery of shRNA targeted against KLHL1 (shKLHL1). Control adenoviruses did not affect any of the neuronal properties measured, yet down-regulation of KLHL1 resulted in HVA current densities ~68% smaller and LVA current densities 44% smaller than uninfected controls, with a concomitant reduction in α_1A and α_1H protein levels. Biophysical analysis and western blot experiments suggest Ca_V3.1 and 3.3 currents are also present in shKLHL1-infected neurons. Synapsin I levels, miniature postsynaptic current frequency, and excitatory and inhibitory synapse number were reduced in KLHL1 knockdown. This study corroborates the physiological role of KLHL1 as a calcium channel modulator and demonstrates a novel, presynaptic role.

Original language	English (US)
Pages (from-to)	269-280
Number of pages	12
Journal	Cell Calcium
Volume	55
Issue number	5
DOIs	https://doi.org/10.1016/j.ceca.2014.03.002
State	Published - May 2014

Bibliographical note

Funding Information:
We thank all members of the Piedras laboratory for helpful comments and discussions. This paper is based upon work supported by the National Science Foundation under Grant no. 1022075 (EPR).

Keywords

ATXN8OS
Actin-binding protein
HVA calcium current
LVA calcium currents
P/Q-type channel
Spinocerebellar ataxia type 8
Synaptic
T-type channel

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title = "Down-regulation of endogenous KLHL1 decreases voltage-gated calcium current density",

abstract = "The actin-binding protein Kelch-like 1 (KLHL1) can modulate voltage-gated calcium channels in vitro. KLHL1 interacts with actin and with the pore-forming subunits of Cav2.1 and CaV3.2 calcium channels, resulting in up-regulation of P/Q and T-type current density. Here we tested whether endogenous KLHL1 modulates voltage gated calcium currents in cultured hippocampal neurons by down-regulating the expression of KLHL1 via adenoviral delivery of shRNA targeted against KLHL1 (shKLHL1). Control adenoviruses did not affect any of the neuronal properties measured, yet down-regulation of KLHL1 resulted in HVA current densities ~68% smaller and LVA current densities 44% smaller than uninfected controls, with a concomitant reduction in α1A and α1H protein levels. Biophysical analysis and western blot experiments suggest CaV3.1 and 3.3 currents are also present in shKLHL1-infected neurons. Synapsin I levels, miniature postsynaptic current frequency, and excitatory and inhibitory synapse number were reduced in KLHL1 knockdown. This study corroborates the physiological role of KLHL1 as a calcium channel modulator and demonstrates a novel, presynaptic role.",

keywords = "ATXN8OS, Actin-binding protein, HVA calcium current, LVA calcium currents, P/Q-type channel, Spinocerebellar ataxia type 8, Synaptic, T-type channel",

author = "Perissinotti, {Paula P.} and Ethington, {Elizabeth G.} and Leanne Cribbs and Koob, {Michael D.} and Jody Martin and Piedras-Renter{\'i}a, {Erika S.}",

note = "Funding Information: We thank all members of the Piedras laboratory for helpful comments and discussions. This paper is based upon work supported by the National Science Foundation under Grant no. 1022075 (EPR). ",

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T1 - Down-regulation of endogenous KLHL1 decreases voltage-gated calcium current density

AU - Perissinotti, Paula P.

AU - Ethington, Elizabeth G.

AU - Cribbs, Leanne

AU - Koob, Michael D.

AU - Martin, Jody

AU - Piedras-Rentería, Erika S.

N1 - Funding Information: We thank all members of the Piedras laboratory for helpful comments and discussions. This paper is based upon work supported by the National Science Foundation under Grant no. 1022075 (EPR).

PY - 2014/5

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N2 - The actin-binding protein Kelch-like 1 (KLHL1) can modulate voltage-gated calcium channels in vitro. KLHL1 interacts with actin and with the pore-forming subunits of Cav2.1 and CaV3.2 calcium channels, resulting in up-regulation of P/Q and T-type current density. Here we tested whether endogenous KLHL1 modulates voltage gated calcium currents in cultured hippocampal neurons by down-regulating the expression of KLHL1 via adenoviral delivery of shRNA targeted against KLHL1 (shKLHL1). Control adenoviruses did not affect any of the neuronal properties measured, yet down-regulation of KLHL1 resulted in HVA current densities ~68% smaller and LVA current densities 44% smaller than uninfected controls, with a concomitant reduction in α1A and α1H protein levels. Biophysical analysis and western blot experiments suggest CaV3.1 and 3.3 currents are also present in shKLHL1-infected neurons. Synapsin I levels, miniature postsynaptic current frequency, and excitatory and inhibitory synapse number were reduced in KLHL1 knockdown. This study corroborates the physiological role of KLHL1 as a calcium channel modulator and demonstrates a novel, presynaptic role.

AB - The actin-binding protein Kelch-like 1 (KLHL1) can modulate voltage-gated calcium channels in vitro. KLHL1 interacts with actin and with the pore-forming subunits of Cav2.1 and CaV3.2 calcium channels, resulting in up-regulation of P/Q and T-type current density. Here we tested whether endogenous KLHL1 modulates voltage gated calcium currents in cultured hippocampal neurons by down-regulating the expression of KLHL1 via adenoviral delivery of shRNA targeted against KLHL1 (shKLHL1). Control adenoviruses did not affect any of the neuronal properties measured, yet down-regulation of KLHL1 resulted in HVA current densities ~68% smaller and LVA current densities 44% smaller than uninfected controls, with a concomitant reduction in α1A and α1H protein levels. Biophysical analysis and western blot experiments suggest CaV3.1 and 3.3 currents are also present in shKLHL1-infected neurons. Synapsin I levels, miniature postsynaptic current frequency, and excitatory and inhibitory synapse number were reduced in KLHL1 knockdown. This study corroborates the physiological role of KLHL1 as a calcium channel modulator and demonstrates a novel, presynaptic role.

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KW - Actin-binding protein

KW - HVA calcium current

KW - LVA calcium currents

KW - P/Q-type channel

KW - Spinocerebellar ataxia type 8

KW - Synaptic

KW - T-type channel

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JO - Cell Calcium

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ER -

Down-regulation of endogenous KLHL1 decreases voltage-gated calcium current density

Abstract

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Keywords

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