The SNARE proteins SNAP-25 and SNAP-23 display different affinities for lipid rafts in PC12 cells. Regulation by distinct cysteine-rich domains

J Biol Chem. 2005 Jan 14;280(2):1236-40. doi: 10.1074/jbc.M410674200. Epub 2004 Nov 12.

Abstract

SNAP-25 and its ubiquitously expressed homologue, SNAP-23, are SNARE proteins that are essential for regulated exocytosis in diverse cell types. Recent work has shown that SNAP-25 and SNAP-23 are partly localized in sphingolipid/cholesterol-rich lipid raft domains of the plasma membrane and that the integrity of these domains is important for exocytosis. Here, we show that raft localization is mediated by a 36-amino-acid region of SNAP-25 that is also the minimal sequence required for membrane targeting; this domain contains 4 closely spaced cysteine residues that are sites for palmitoylation. Analysis of endogenous levels of SNAP-25 and SNAP-23 present in lipid rafts in PC12 cells revealed that SNAP-23 (54% raft-associated) was almost 3-fold more enriched in rafts when compared with SNAP-25 (20% raft-associated). We report that the increased raft association of SNAP-23 occurs due to the substitution of a highly conserved phenylalanine residue present in SNAP-25 with a cysteine residue. Intriguingly, although the extra cysteine in SNAP-23 enhances its raft association, the phenylalanine at the same position in SNAP-25 acts to repress the raft association of this protein. These different raft-targeting signals within SNAP-25 and SNAP-23 are likely important for fine-tuning the exocytic pathways in which these proteins operate.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Binding Sites
  • Carrier Proteins / genetics
  • Carrier Proteins / metabolism*
  • Cysteine / genetics
  • Cysteine / metabolism
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism
  • Membrane Microdomains / metabolism*
  • Membrane Proteins / genetics
  • Membrane Proteins / metabolism*
  • Mice
  • Molecular Sequence Data
  • Mutation / genetics
  • Nerve Tissue Proteins / genetics
  • Nerve Tissue Proteins / metabolism*
  • PC12 Cells
  • Palmitic Acid / metabolism
  • Phenylalanine / genetics
  • Phenylalanine / metabolism
  • Protein Structure, Tertiary
  • Protein Transport
  • Qb-SNARE Proteins
  • Qc-SNARE Proteins
  • Rats
  • SNARE Proteins
  • Synaptosomal-Associated Protein 25
  • Vesicular Transport Proteins / genetics
  • Vesicular Transport Proteins / metabolism*

Substances

  • Carrier Proteins
  • Membrane Proteins
  • Nerve Tissue Proteins
  • Qb-SNARE Proteins
  • Qc-SNARE Proteins
  • SNAP23 protein, human
  • SNARE Proteins
  • Snap23 protein, mouse
  • Snap25 protein, mouse
  • Snap25 protein, rat
  • Synaptosomal-Associated Protein 25
  • Vesicular Transport Proteins
  • Green Fluorescent Proteins
  • Palmitic Acid
  • Phenylalanine
  • Cysteine