Munc18: A Presynaptic Transmitter Release Site N type (CaV2.2) Calcium Channel Interacting Protein

• Published in: Channels (Austin, Tex.) Vol. 1; no. 1; pp. 12 - 21
• Main Authors: Chan, Allen W., Khanna, Rajesh, Li, Qi, Stanley, Elise F.
• Format: Journal Article
• Language: English
• Published: United States Taylor & Francis 01-01-2007
• Subjects: Alternative Splicing - physiology; Animals; Base Sequence; Binding; Biology; Bioscience; Calcium; Calcium Channels, N-Type - genetics; Calcium Channels, N-Type - metabolism; Cancer; Cell; Cell Line; Chick Embryo; Chickens; Cycle; Ganglia, Spinal - cytology; Ganglia, Spinal - metabolism; Humans; Kinetics; Landes; Molecular Sequence Data; Munc18 Proteins - genetics; Munc18 Proteins - metabolism; Nerve Tissue Proteins - genetics; Nerve Tissue Proteins - metabolism; Neurotransmitter Agents - genetics; Neurotransmitter Agents - metabolism; Organogenesis; Presynaptic Terminals - metabolism; Protein Binding - physiology; Protein Structure, Tertiary - physiology; Proteins; Rats; Rats, Sprague-Dawley; SNARE Proteins - genetics; SNARE Proteins - metabolism; Synaptic Vesicles - genetics; Synaptic Vesicles - metabolism; Synaptotagmins - genetics; Synaptotagmins - metabolism; Syntaxin 1 - genetics; Syntaxin 1 - metabolism
• ISSN: 1933-6950; 1933-6969
• DOI: 10.4161/chan.3694

Munc18 is a presynaptic protein that is essential for transmitter release. Recent studies have indicated that this protein is involved in secretory vesicle docking but its binding partners in this role remain a mystery. We demonstrate using the isolated calyx-type presynaptic terminal of the chick ciliary ganglion that staining for Munc18 co-localizes and covaries with that for transmitter release site N type calcium channels (CaV2.2), consistent with elements of a common release site complex. Biochemical analysis demonstrated that the protein co-precipitates with CaV2.2 from lysates of rat or chick brain, including the synaptic, long-splice variant; presynaptic terminal surface membrane proteins, and a cell line co-expressing Munc18 and CaV2.2. Munc18 bound with high affinity to the CaV2.2 II-III intracellular loop, low affinity to the I-II loop but not to other channel intracellular regions. Over-expression of Munc18 in dorsal root ganglion neurons did not affect CaV2.2 current amplitude or fast kinetics but siRNA-knockdown resulted in a negative shift in the steady state inactivation curve, an effect attributed to an indirect action via syntaxin 1. Recombinant Munc18 also co-precipitated strongly with the v-SNARE synaptotagmin but only weakly with other SNAREs. Thus, the calcium channel may serve as a surface membrane platform anchoring a Munc18-containing bridge to synaptotagmin and the synaptic vesicle.