Involvement of annexin II in exocytosis of lamellar bodies from alveolar epithelial type II cells

Annexins are a family of Ca2+- and phospholipid-binding proteins that have been implicated in exocytosis. In the present study, we investigated the participation of selected annexins in exocytosis of lamellar bodies by examining their liposome aggregation property and ability to reconstitute surfactant secretion from permeabilized rat lung alveolar type II cells. Annexins I, II, III, and VI were demonstrated in type II cells by immunoblot analysis, but annexin IV and V were not found. Annexins I-IV mediated liposome aggregation in the presence of 1 mM Ca2+. However, only annexin II tetramer had aggregation activity at 10 mu M Ca2+. Annexins V and VI had negligible aggregation activity at any Ca2+ concentrations (up to 1 mM Ca2+). To study reconstitution of secretion by annexins, isolated type II cells were permeabilized with 40 mu M beta-escin. Under these conditions, the permeabilized cells released similar to 30-40% lactic acid dehydrogenase into the medium. An undetermined fraction of cellular annexin content was lost during permeabilization. However, lamellar bodies in the permeabilized type II cells stained appropriately with the fluorescent dyes Nile red and quinacrine, indicating that they were intact. These permeabilized cells were secretion competent, since phosphatidylcholine (PC) secretion was stimulated by 0.2-1.0 mu M Ca2+. Addition of an exogenous annexin mixture enhanced PC secretion from the permeabilized type IV cells with maximal stimulation at 0.5 mu M Ca2+. Of six purified annexins (I-VI) tested for their ability to reconstitute secretion from the permeabilized cells, only annexin II was effective. Our results suggest that annexin II is necessary for exocytosis of lamellar bodies.