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 investiga ted the participation of selected annexins in exocytosis of lamellar b odies 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 Ca 2+. However, only annexin II tetramer had aggregation activity at 10 m u M Ca2+. Annexins V and VI had negligible aggregation activity at any Ca2+ concentrations (up to 1 mM Ca2+). To study reconstitution of sec retion by annexins, isolated type II cells were permeabilized with 40 mu M beta-escin. Under these conditions, the permeabilized cells relea sed similar to 30-40% lactic acid dehydrogenase into the medium. An un determined fraction of cellular annexin content was lost during permea bilization. However, lamellar bodies in the permeabilized type II cell s stained appropriately with the fluorescent dyes Nile red and quinacr ine, indicating that they were intact. These permeabilized cells were secretion competent, since phosphatidylcholine (PC) secretion was stim ulated by 0.2-1.0 mu M Ca2+. Addition of an exogenous annexin mixture enhanced PC secretion from the permeabilized type IV cells with maxima l stimulation at 0.5 mu M Ca2+. Of six purified annexins (I-VI) tested for their ability to reconstitute secretion from the permeabilized ce lls, only annexin II was effective. Our results suggest that annexin I I is necessary for exocytosis of lamellar bodies.