Polarized secretory epithelial cells possess several sorting devices to effect vectorial flow of secretory and membrane proteins to their apical and basolateral poles. Our previous studies on pancreatic acinar cells have delineated the kinetics and routes of intracellular transpost of secretory proteins that enter a secretagogue activated (regulated) pathway leading to apical exocytosis while a separate secretory pathway results in basolateral, nonstimulated (constitutive) discharge of basement membrane proteins. Similarly, vectorial delivery of membrane proteins to the apical and basolateral poles of secretory cells is required to establish and maintain structural and functional poolarity. The goal of this project is to define the routes and control mechanisms for constitutive and regulated secretory pathways in pancreatic acinar and other epithelial cells and to characterize the pathways responsible for biogenesis of plasmalemmal polarity. To this end, we will use pancreatic acinar and other cell lines cultured in chambers that allow probing of apical and basolateral secretory compartments. Agents that perturb secretory protein sorting will be tested for their ability to affect differentially apical (e.g. amylase) or basolateral (e.g. basal lamina) secretory pathways. These will include secretagogues, acidothrophic agents, inhibitors of glycosylation and protein synthesis and drugs producing cytoskeletal disassembly. The effects of reduction in temperature and ATP levels will also be examined. Comparable experimental conditions will be used, in conjunction with immnoassays for polarity of apical (gamma-glutamyl transferase) and basolateral (insulin and laminin receptors) membrane protein delivery, to define factors regulating biogenesis of plasmalemmal polarity and to clarify relationships between secretory protein and membrane protein delivery pathways. We will also examine membrane targeting in MDCX cells transfected with cDNAs for gamma-glutamyl transferase, an apical and basolateral membrane protein in pancreatic acinar cells. Finally, we plan to study biogenesis of the regulated secretory pathway in developing pancreas as it acquires secretagogue responsiveness.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37DK017389-17
Application #
3483201
Study Section
Cellular Biology and Physiology Subcommittee 1 (CBY)
Project Start
1977-08-01
Project End
1992-07-31
Budget Start
1989-08-01
Budget End
1990-07-31
Support Year
17
Fiscal Year
1989
Total Cost
Indirect Cost
Name
Yale University
Department
Type
Schools of Medicine
DUNS #
082359691
City
New Haven
State
CT
Country
United States
Zip Code
06520
Valentijn, J A; Valentijn, K; Pastore, L M et al. (2000) Actin coating of secretory granules during regulated exocytosis correlates with the release of rab3D. Proc Natl Acad Sci U S A 97:1091-5
Valentijn, K M; Gumkowski, F D; Jamieson, J D (1999) The subapical actin cytoskeleton regulates secretion and membrane retrieval in pancreatic acinar cells. J Cell Sci 112 ( Pt 1):81-96
Valentijn, J A; Jamieson, J D (1998) Carboxyl methylation of rab3D is developmentally regulated in the rat pancreas: correlation with exocrine function. Eur J Cell Biol 76:204-11
Valentijn, J A; LaCivita, D Q; Gumkowski, F D et al. (1997) Rab4 associates with the actin terminal web in developing rat pancreatic acinar cells. Eur J Cell Biol 72:1-8
Valentijn, J A; Sengupta, D; Gumkowski, F D et al. (1996) Rab3D localizes to secretory granules in rat pancreatic acinar cells. Eur J Cell Biol 70:33-41
Valentijn, J A; Gumkowski, F D; Jamieson, J D (1996) The expression pattern of rab3D in the developing rat exocrine pancreas coincides with the acquisition of regulated exocytosis. Eur J Cell Biol 71:129-36
Sengupta, D; Gumkowski, F D; Tang, L H et al. (1996) Localization of cellubrevin to the Golgi complex in pancreatic acinar cells. Eur J Cell Biol 70:306-14
Jena, B P; Gumkowski, F D; Konieczko, E M et al. (1994) Redistribution of a rab3-like GTP-binding protein from secretory granules to the Golgi complex in pancreatic acinar cells during regulated exocytosis. J Cell Biol 124:43-53
O'Sullivan, A J; Jamieson, J D (1992) Activation of protein kinase C is not an absolute requirement for amylase release from permeabilized rat pancreatic acini. Biochem J 285 ( Pt 2):597-601
Nathanson, M H; Padfield, P J; O'Sullivan, A J et al. (1992) Mechanism of Ca2+ wave propagation in pancreatic acinar cells. J Biol Chem 267:18118-21

Showing the most recent 10 out of 22 publications