Calcium-dependence of synexin binding may determine aggregation and fusion of lamellar bodies.
Synexin (annexin VII) is a member of the annexin family of calcium and phospholipid binding proteins that promote calcium-dependent aggregation and fusion of lipid vesicles or secretory granules. We have previously suggested that synexin may be involved in membrane fusion processes during exocytosis of lung surfactant since it promotes fusion in vitro of lamellar bodies with plasma membranes. In this study, we characterized calcium-dependency of synexin binding to lamellar bodies and plasma membranes, since such binding is the initial, and, therefore, may be the rate-limiting step in membrane aggregation and fusion. The binding of biotinylated synexin to lamellar bodies and plasma membranes increased in a calcium-dependent manner reaching a maximum at approx. 200 microM Ca2+. Binding to lamellar bodies was completely inhibited by unlabelled synexin. Gel-overlay analysis showed that synexin bound to an approx. 76 kDa protein in the lamellar body and plasma membrane fractions. The calcium kinetics were noticeably similar for synexin binding to lamellar bodies and plasma membranes, aggregation of lamellar bodies, and fusion of lamellar bodies with lipid vesicles. At low calcium concentrations, aggregation of lamellar bodies could be increased with increasing synexin concentration, and arachidonic acid increased all three parameters (binding, aggregation, and fusion) in a similar manner. The effects of calcium and arachidonic acid on these three parameters suggest that synexin binding to lamellar bodies may be a rate-determining step for fusion during surfactant secretion. Furthermore, at near physiological calcium levels, the membrane fusion may be enhanced by elevated concentrations of synexin and polyunsaturated fatty acids.
Sen
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N
Department of Pediatrics, Jefferson Medical College, Thomas Jefferson University, Philadelphia, PA 19107, USA.
Spitzer
A R
AR
Chander
A
A
D000818:Animals; D017310:Annexin A7 / Q000378:metabolism* / Q000494:pharmacology; D001667:Binding, Competitive; D002118:Calcium / Q000502:physiology*; D002417:Cattle; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology; D007425:Intracellular Membranes / Q000187:drug effects / Q000502:physiology; D008168:Lung; D008561:Membrane Fusion / Q000187:drug effects / Q000502:physiology*; D015388:Organelles / Q000187:drug effects / Q000502:physiology*; D011485:Protein Binding / Q000187:drug effects; D051381:Rats
D017310:Annexin A7; D002118:Calcium