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R-type voltage-gated Ca(2+) channel interacts with synaptic proteins and recruits synaptotagmin to the plasma membrane of Xenopus oocytes.

Abstract：

:It is well established that syntaxin 1A, synaptosomal-associated protein of 25 kDa (SNAP-25) and synaptotagmin either alone or in combination, modulate the kinetic properties of voltage-gated Ca(2+) channels Ca(v)1.2 (Lc-channel) Ca(v)2.2 (N-type) and Ca(v)2.1 (P/Q-type). The interaction interface was found to reside at the cytosolic II-III domain of the alpha1 subunit of the channels. In this study, we demonstrated a functional coupling of human neuronal Ca(v)2.3 (R-type channel) with syntaxin 1A, SNAP-25 and synaptotagmin in BAPTA injected Xenopus oocytes. The kinetic properties of Ca(v)2.3 assembled with syntaxin 1A, SNAP-25 or synaptotagmin individually differed from Ca(v)2.3 associated with binary complexes syntaxin 1A/SNAP-25, syntaxin 1A/synaptotagmin or SNAP-25/synaptotagmin. Co-expression of Ca(v)2.3 with syntaxin 1A, SNAP-25 and synaptotagmin together, produced a channel with distinctive kinetic properties analogous to excitosome multiprotein complex generated by Ca(v)1.2 and Ca(v)2.2. Exchanging the current-carrying ions altered the kinetics of channel/synaptic proteins interaction, indicating a tight crosstalk formed between the permeation pathway of Ca(v)2.3 and the fusion apparatus during membrane depolarization. This putative coupling could predict how the release site might be organized to allow a rapid communication between the channel and the release machinery. In vivo confocal imaging of oocytes revealed GFP-synaptotagmin at the plasma membrane when the channel was present, as opposed to random distribution in its absence, consistent with Ca(2+)-independent molecular link of synaptotagmin and the channel. Synaptotagmin was detected at the membrane also in oocytes co-expressing the soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs). Both imaging studies and protein-protein interactions in Xenopus oocytes show that channel linkage to synaptotagmin precedes Ca(2+) influx. Altogether, the R-type channel appears to associate with synaptic proteins to generate a multiprotein excitosome complex prior to Ca(2+)-entry. We propose that the distinct kinetics of the Ca(2+)-channel acquired by the close association with the vesicle and the t-SNAREs within the excitosome complex may be essential for depolarization evoked transmitter release.

journal_name

Neuroscience

journal_title

Neuroscience

authors

Cohen R,Atlas D

doi

10.1016/j.neuroscience.2004.07.027

keywords:

subject

Has Abstract

pub_date

2004-01-01 00:00:00

pages

831-41

issue

eissn

0306-4522

issn

1873-7544

pii

S0306-4522(04)00592-5

journal_volume

128

pub_type

杂志文章

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R-type voltage-gated Ca(2+) channel interacts with synaptic proteins and recruits synaptotagmin to the plasma membrane of Xenopus oocytes.