Abstract
Intracellular membrane fusion is mediated by the concerted action of N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) and Sec1/Munc18 (SM) proteins. During fusion, SM proteins bind the N-terminal peptide (N-peptide) motif of the SNARE subunit syntaxin, but the function of this interaction is unknown. Here, using FRET-based biochemical reconstitution and Caenorhabditis elegans genetics, we show that the N-peptide of syntaxin-1 recruits the SM protein Munc18-1/nSec1 to the SNARE bundle, facilitating their assembly into a fusion-competent complex. The recruitment is achieved through physical tethering rather than allosteric activation of Munc18-1. Consistent with the recruitment role, the N-peptide is not spatially constrained along syntaxin-1, and it is functional when translocated to another SNARE subunit SNAP-25 or even when simply anchored in the target membrane. The N-peptide function is restricted to an early initiation stage of the fusion reaction. After association, Munc18-1 and the SNARE bundle together drive membrane merging without further involving the N-peptide. Thus, the syntaxin N-peptide is an initiation factor for the assembly of the SNARE-SM membrane fusion complex.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
MeSH terms
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Amino Acid Motifs / genetics
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Amino Acid Sequence
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Animals
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Caenorhabditis elegans / genetics
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Caenorhabditis elegans / metabolism
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Caenorhabditis elegans Proteins / chemistry
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Caenorhabditis elegans Proteins / genetics
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Caenorhabditis elegans Proteins / metabolism*
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Female
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Green Fluorescent Proteins / genetics
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Green Fluorescent Proteins / metabolism
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Male
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Membrane Fusion*
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Microscopy, Confocal
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Models, Molecular
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Multiprotein Complexes / chemistry
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Multiprotein Complexes / metabolism
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Munc18 Proteins / chemistry
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Munc18 Proteins / genetics
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Munc18 Proteins / metabolism
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Mutation
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Phosphoproteins / chemistry
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Phosphoproteins / genetics
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Phosphoproteins / metabolism*
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Protein Binding
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Protein Conformation
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Protein Structure, Tertiary
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SNARE Proteins / chemistry
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SNARE Proteins / genetics
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SNARE Proteins / metabolism*
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Syntaxin 1 / chemistry
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Syntaxin 1 / genetics
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Syntaxin 1 / metabolism*
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Time Factors
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Vesicular Transport Proteins / chemistry
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Vesicular Transport Proteins / genetics
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Vesicular Transport Proteins / metabolism*
Substances
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Caenorhabditis elegans Proteins
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Multiprotein Complexes
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Munc18 Proteins
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Phosphoproteins
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SNARE Proteins
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Syntaxin 1
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Unc-18 protein, C elegans
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Vesicular Transport Proteins
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Green Fluorescent Proteins