Regulation of syntaxin1A-munc18 Complex for SNARE Pairing in HEK293 Cells

J Physiol. 2004 Aug 1;558(Pt 3):857-71. doi: 10.1113/jphysiol.2004.067249. Epub 2004 Jun 24.

Abstract

The formation and dissolution of SNARE protein complexes is essential for Ca(2+)-triggered fusion of neurotransmitter-filled vesicles at the presynaptic membrane. Among the pre-synaptic SNARE proteins, the activation of the Q-SNARE syntaxin1A is a critical event for SNARE complex formation. Activation requires syntaxin1A to transit from a munc18-bound non-interacting state to one competent for SNARE binding. The molecular mechanisms that regulate this transition remain unclear. The propensity of syntaxin1A to promote voltage-dependent steady-state inactivation of N-type Ca(2+) channels and accelerate their entry into inactivation was used in a heterologous cell expression system to elucidate regulation of syntaxin1A protein-protein interactions. We report that coexpression of munc18 eliminated the promoting effect of syntaxin1A on inactivation. This effect of munc18 was completely disrupted by coexpression of munc13-1, but not munc13-2 or munc13-3. Also, since expression of munc13-1 with syntaxin1A resulted in an inactivation phenotype identical to that of munc18 with syntaxin1A, the action of munc13-1 on the munc18-syntaxin1A complex was functionally unique and did not result from competitive binding interactions. Furthermore, munc13 expressed with syntaxin1A and munc18 promoted redistribution of a cytosolic SNAP25 mutant to the membrane, a result indicative of syntaxin1A-SNAP25 SNARE pairing. These data demonstrate an important role of munc13 to control the protein-protein interactions of syntaxin1A in vivo, and support munc13 as critical to dissociating syntaxin1A-munc18 complexes and making syntaxin1A available for SNARE interactions.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Antigens, Surface / biosynthesis*
  • Antigens, Surface / genetics
  • Cell Line
  • Humans
  • Membrane Potentials / physiology
  • Munc18 Proteins
  • Nerve Tissue Proteins / biosynthesis*
  • Nerve Tissue Proteins / genetics
  • Protein Binding / physiology
  • Rats
  • SNARE Proteins
  • Syntaxin 1
  • Vesicular Transport Proteins / biosynthesis*
  • Vesicular Transport Proteins / genetics

Substances

  • Antigens, Surface
  • Munc18 Proteins
  • Nerve Tissue Proteins
  • SNARE Proteins
  • Syntaxin 1
  • Vesicular Transport Proteins