Pulmonary surfactant secreted by type II pneumocytes facilitates pulmonary expansion at birth and prevents the alveolar atelectasis found in both the neonatal and adult forms of respiratory distress rome. There is neither a large intracellular nor extracellular reserve of pulmonary surfactant. We will investigate specific molecular phosphorylation and dephosphorylation mechanisms which regulate surfactant secretion in rat type II pneumocytes in culture. Surfactant secretion is stimulated by beta and Pl receptors which increase CAMP production and CAMP-dependent protein kinase activation, P2 receptors which stimulate IP formation and intracellular calcium release, and by phorbol esters which stimulate protein kinase C.
The Specific Aims of this project are: 1. to map, identify and compare target proteins which are phosphorylated """"""""in vivo"""""""" and in vitro in rat type II pneumocytes by activation of CAMP-dependent protein kinase or protein kinase C and Proline-Directed Protein Kinase, a novel growth factor activated S/T kinase; 2. to assess the functional effects of phosphorylation in these proteins by correlation with surfactant secretion rate, and F-actin polymerization; 3. to identify the protein phosphatases which are active in type II pneumocytes in culture, and to investigate their role in the regulation of protein phosphorylation and surfactant secretion using kadaic acid, a specific inhibitor; 4. to define the ontogeny of protein phosphorylation in fetal, neonatal and adult cells. These studies will characterize specific sites of molecular regulation of surfactant secretion mediated protein phosphorylation and dephosphorylation in type II pneumocytes. This will lead to novel molecular strategies for the prevention and treatment of both the neonatal and adult forms of respiratory distress syndrome.
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