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Mol Pharmacol. 2000 Mar;57(3):519-28.

Calcium-independent receptor for alpha-latrotoxin and neurexin 1alpha [corrected] facilitate toxin-induced channel formation: evidence that channel formation results from tethering of toxin to membrane.

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  • 1Department of Pharmacology, University of Michigan Medical School, Ann Arbor, Michigan, USA.

Erratum in

  • Mol Pharmacol 2000 Apr;57(4):832.

Abstract

alpha-Latrotoxin binding to the calcium-independent receptor for alpha-latrotoxin (CIRL-1), a putative G-protein-coupled receptor, stimulates secretion from chromaffin and PC12 cells. Using patch clamp techniques and microspectrofluorimetry, we demonstrate that the interaction of alpha-latrotoxin with CIRL-1 produces a high conductance channel that permits increases in cytosolic Ca(2+). alpha-Latrotoxin interaction with CIRL-1 transiently expressed in bovine chromaffin cells produced a 400-pS channel, which rarely closed under Ca(2+)-free conditions. The major effect of overexpressing CIRL-1 was to greatly increase the sensitivity of chromaffin cells to channel formation by alpha-latrotoxin. alpha-Latrotoxin interaction with CIRL-1 transiently overexpressed in non-neuronal human embryonic kidney 293 (HEK293) cells produced channels that were nearly identical with those observed in chromaffin cells. Channel currents were reduced by millimolar Ca(2+). At alpha-latrotoxin concentrations below 500 pM, channel formation occurred many seconds after binding of toxin to CIRL-1 indicating distinct steps in channel formation. In all cases there was a rapid, sequential addition of channels once the first channel appeared. An analysis of CIRL-1 mutants indicated that channel formation in HEK293 cells is unlikely to be transduced by a G-protein-dependent mechanism. alpha-Latrotoxin interaction with a fusion construct composed of the extracellular domain of CIRL-1 anchored to the membrane by the transmembrane domain of vesicular stomatitis virus glycoprotein, and with neurexin 1alpha, an alpha-latrotoxin receptor structurally unrelated to CIRL-1, produced channels virtually identical with those observed with wild-type CIRL-1. We propose that alpha-latrotoxin receptors recruit toxin to facilitate its insertion across the membrane and that alpha-latrotoxin itself controls the conductance properties of the channels it produces.

PMID:
10692492
[PubMed - indexed for MEDLINE]
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