title: Regulation of syntaxin1A-munc18 complex for SNARE pairing in HEK293 cells.
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.
authors.affiliation: 7804 Medical Sciences II Building, Department of Molecular and Integrative Physiology, The Medical School, University of Michigan, Ann Arbor, MI 48109-0622, USA. segladys@umich.edu
authors.forename: Svetlana E
authors.initials: SE
authors.lastname: Gladycheva
authors.forename: Chi S
authors.initials: CS
authors.lastname: Ho
authors.forename: Yue Ying F
authors.initials: YY
authors.lastname: Lee
authors.forename: Edward L
authors.initials: EL
authors.lastname: Stuenkel
mesh_terms: D000818:Animals; D000954:Antigens, Surface / Q000096:biosynthesis* / Q000235:genetics; D002460:Cell Line; D006801:Humans; D008564:Membrane Potentials / Q000502:physiology; D051938:Munc18 Proteins; D009419:Nerve Tissue Proteins / Q000096:biosynthesis* / Q000235:genetics; D011485:Protein Binding / Q000502:physiology; D051381:Rats; D050600:SNARE Proteins; D050827:Syntaxin 1; D033921:Vesicular Transport Proteins / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D000954:Antigens, Surface; D051938:Munc18 Proteins; D009419:Nerve Tissue Proteins; D050600:SNARE Proteins; D050827:Syntaxin 1; D033921:Vesicular Transport Proteins