Physical and functional interactions of SNAP-23 with annexin A2

Am J Respir Cell Mol Biol. 2007 Oct;37(4):467-76. doi: 10.1165/rcmb.2006-0447OC. Epub 2007 Jun 15.

Abstract

Lung surfactant is secreted through the fusion of lamellar bodies with the plasma membrane of alveolar epithelial type II cells. Annexin A2, a Ca(2+)- and phospholipid-binding protein, promotes the fusion of lamellar bodies with the plasma membrane. Soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) are known to have an essential role in surfactant secretion. We hypothesized that annexin A2 acts as a Ca(2+) sensor and mediates membrane fusion via its interaction with SNAREs. Both purified or endogenous annexin A2 in type II cells specifically bound with SNAP-23 in a Ca(2+)-dependent manner, as determined by pull-down experiments using recombinant glutathione S-transferase-tagged SNAP-23. A deletion study identified the cysteine-rich region (CRR) of SNAP-23 as the binding site for annexin A2. Mutations of cysteine residues in the CRR dramatically decreased the binding. SNAP-23 also co-immunoprecipitated with annexin A2; however, a SNAP-23 mutant failed to co-immunoprecipitate with annexin A2. Immunofluorescence revealed a co-localization of SNAP-23 and annexin A2 in type II cells. Furthermore, anti-SNAP-23 antibody significantly inhibited annexin A2-mediated fusion between lamellar bodies and the plasma membrane. These data suggest that annexin A2 and SNAP-23 are involved in the same pathway in the regulation of lung surfactant secretion.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Annexin A2 / isolation & purification
  • Annexin A2 / metabolism*
  • Antibodies / pharmacology
  • Binding Sites
  • Cattle
  • Cell Membrane / drug effects
  • Cell Membrane / metabolism
  • Cysteine
  • Humans
  • Immunoprecipitation
  • Mammals
  • Molecular Sequence Data
  • Mutant Proteins / metabolism
  • Protein Binding / drug effects
  • Protein Isoforms / metabolism
  • Pulmonary Alveoli / cytology
  • Pulmonary Alveoli / drug effects
  • Pulmonary Alveoli / metabolism
  • Rats
  • Rats, Sprague-Dawley
  • Sequence Deletion
  • Two-Hybrid System Techniques
  • Vesicular Transport Proteins / chemistry
  • Vesicular Transport Proteins / metabolism*

Substances

  • Annexin A2
  • Antibodies
  • Mutant Proteins
  • Protein Isoforms
  • Snap23 protein, rat
  • Vesicular Transport Proteins
  • Cysteine