Novel isoform of syntaxin 1 is expressed in mammalian cells.
Syntaxin 1A has been identified previously as a neural-cell-specific, membrane-anchored receptor protein required for docking and fusion of synaptic vesicles with the presynaptic plasma membrane. Syntaxin 1A consists of 288 amino acid residues including a 265-residue N-terminal region exposed to the cytoplasm and a C-terminal hydrophobic stretch of 23 residues believed to anchor syntaxin to the plasma membrane. Using a human fat-cell library we have isolated a novel cDNA clone of syntaxin 1A containing an insert of 91 bp in codon 226. This insert and subsequent frame shift generated a cDNA that codes for a truncated protein of 260 residues without the C-terminal transmembrane domain characteristic of the syntaxin family. Analysis of the deduced amino acid sequence of the new cDNA clone, termed syntaxin 1C, showed that it was identical for the first 226 residues with the previously described neural syntaxin 1A, and diverged thereafter. The truncated protein lacked the botulinum neurotoxin C cleavage site (Lys253-Ala254), a feature of the syntaxin 1A protein, because of the novel C-terminal domain of 34 residues. The new C-terminal region contained a single cysteine residue and was moderately rich in proline, with three repeats of a PXP motif. The insert occurred within the region encoding the coiled-coil motifs required for interactions with synaptobrevin, alpha-SNAP (SNAP being soluble N-ethylmaleimide-sensitive factor attachment protein) and n-Sec1/Munc-18 (n-Sec1 being the rat brain homologue of yeast Sec1p and Munc-18 the mammalian homologue of Caenorhabditis elegans unc-18, but five residues outside the domain previously mapped as being required for binding SNAP-25. Interaction studies in vitro suggested that unlike syntaxin 1A, which binds to both Munc-18a and- 18b, syntaxin 1C binds only to Munc-18b. The new isoform syntaxin 1C, which might be generated by alternative splicing of the syntaxin 1 gene, was expressed in several human tissues, including brain. Immuno-precipitation and immunoblotting with the monoclonal antibody HPC-1 and a polyclonal antibody raised against a peptide corresponding to the unique C-terminal 35 residues of syntaxin 1C failed to detect syntaxin 1C at the protein level in extracts of muscle, fat or brain.
Jagadish
M N
MN
CSIRO, Division of Biomolecular Engineering, Parkville, Victoria, Australia.
Tellam
J T
JT
Macaulay
S L
SL
Gough
K H
KH
James
D E
DE
Ward
C W
CW
D000595:Amino Acid Sequence; D000818:Animals; D000954:Antigens, Surface / Q000737:chemistry*; D001483:Base Sequence; D015139:Blotting, Southern; D001923:Brain Chemistry; D003001:Cloning, Molecular; D006801:Humans; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000737:chemistry*; D016133:Polymerase Chain Reaction; D051381:Rats; D050827:Syntaxin 1
D000954:Antigens, Surface; D009419:Nerve Tissue Proteins; C494212:STX1A protein, human; C494213:Stx1a protein, rat; D050827:Syntaxin 1