Munc13-4 reconstitutes calcium-dependent SNARE-mediated membrane fusion

J Cell Biol. 2012 Apr 16;197(2):301-12. doi: 10.1083/jcb.201109132.

Abstract

Munc13-4 is a widely expressed member of the CAPS/Munc13 protein family proposed to function in priming secretory granules for exocytosis. Munc13-4 contains N- and C-terminal C2 domains (C2A and C2B) predicted to bind Ca(2+), but Ca(2+)-dependent regulation of Munc13-4 activity has not been described. The C2 domains bracket a predicted SNARE-binding domain, but whether Munc13-4 interacts with SNARE proteins is unknown. We report that Munc13-4 bound Ca(2+) and restored Ca(2+)-dependent granule exocytosis to permeable cells (platelets, mast, and neuroendocrine cells) dependent on putative Ca(2+)-binding residues in C2A and C2B. Munc13-4 exhibited Ca(2+)-stimulated SNARE interactions dependent on C2A and Ca(2+)-dependent membrane binding dependent on C2B. In an apparent coupling of membrane and SNARE binding, Munc13-4 stimulated SNARE-dependent liposome fusion dependent on putative Ca(2+)-binding residues in both C2A and C2B domains. Munc13-4 is the first priming factor shown to promote Ca(2+)-dependent SNARE complex formation and SNARE-mediated liposome fusion. These properties of Munc13-4 suggest its function as a Ca(2+) sensor at rate-limiting priming steps in granule exocytosis.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Blood Platelets / metabolism
  • Calcium / metabolism*
  • Calcium-Binding Proteins / metabolism
  • Exocytosis / physiology
  • Humans
  • Liposomes / metabolism
  • Mast Cells / metabolism
  • Membrane Fusion / physiology*
  • Membrane Proteins / metabolism*
  • Neuroendocrine Cells / metabolism
  • SNARE Proteins / metabolism*
  • Synaptotagmins / metabolism

Substances

  • Calcium-Binding Proteins
  • Liposomes
  • Membrane Proteins
  • SNARE Proteins
  • UNC13D protein, human
  • Synaptotagmins
  • Calcium