A syntaxin 1, Galpha(o), and N-type calcium channel complex at a presynaptic nerve terminal: analysis by quantitative immunocolocalization.

Abstract

Presynaptic Ca(V)2.2 (N-type) calcium channels are subject to modulation by interaction with syntaxin 1 and by a syntaxin 1-sensitive Galpha(O) G-protein pathway. We used biochemical analysis of neuronal tissue lysates and a new quantitative test of colocalization by intensity correlation analysis at the giant calyx-type presynaptic terminal of the chick ciliary ganglion to explore the association of Ca(V)2.2 with syntaxin 1 and Galpha(O). Ca(V)2.2 could be localized by immunocytochemistry (antibody Ab571) in puncta on the release site aspect of the presynaptic terminal and close to synaptic vesicle clouds. Syntaxin 1 coimmunoprecipitated with Ca(V)2.2 from chick brain and chick ciliary ganglia and was widely distributed on the presynaptic terminal membrane. A fraction of the total syntaxin 1 colocalized with the Ca(V)2.2 puncta, whereas the bulk colocalized with MUNC18-1. Galpha(O,) whether in its trimeric or monomeric state, did not coimmunoprecipitate with Ca(V)2.2, MUNC18-1, or syntaxin 1. However, the G-protein exhibited a punctate staining on the calyx membrane with an intensity that varied in synchrony with that for both Ca channels and syntaxin 1 but only weakly with MUNC18-1. Thus, syntaxin 1 appears to be a component of two separate complexes at the presynaptic terminal, a minor one at the transmitter release site with Ca(V)2.2 and Galpha(O), as well as in large clusters remote from the release site with MUNC18-1. These syntaxin 1 protein complexes may play distinct roles in presynaptic biology.

• Authors: Qi Li, Anthony Lau, Terence J Morris, Lin Guo, Christopher B Fordyce, Elise F Stanley
• Journal: The Journal of neuroscience : the official journal of the Society for Neuroscience (J Neurosci) Vol. 24 Issue 16 Pg. 4070-81 (Apr 21 2004) ISSN: 1529-2401 [Electronic] United States
• PMID: 15102922 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Antigens, Surface
• Calcium Channels, N-Type
• Macromolecular Substances
• Munc18 Proteins
• Nerve Tissue Proteins
• Stx1a protein, rat
• Stxbp1 protein, rat
• Syntaxin 1
• Vesicular Transport Proteins
• GTP-Binding Protein alpha Subunits, Gi-Go

Topics

• Animals
• Antibody Specificity
• Antigens, Surface (chemistry, metabolism)
• Blotting, Western
• Brain Chemistry
• Calcium Channels, N-Type (chemistry, metabolism)
• Cells, Cultured
• Chick Embryo
• GTP-Binding Protein alpha Subunits, Gi-Go (chemistry, metabolism)
• Immunohistochemistry (methods)
• Macromolecular Substances
• Munc18 Proteins
• Nerve Tissue Proteins (chemistry, metabolism)
• Presynaptic Terminals (metabolism)
• Rats
• Syntaxin 1
• Vesicular Transport Proteins (metabolism)