The long-term objective of the research proposed in this application is to provide new insights into the mechanisms of salivary protein secretion. The intracellular events that couple the input of secretory stimuli and generation of second messengers to the discharge of salivary proteins are largely unknown but are likely to be mediated by proteins of the secretion granule and plasma membranes. The focus of these studies is on the structure and function of certain polypeptides that are components of the membranes of the secretion granules of parotid acinar cells. These polypeptides have been identified using monoclonal antibodies and are among the first to be reported as common components of storage granule membranes in exocrine, endocrine and neural cells. Their broad distribution suggests that they may be common denominators in the distal steps of protein export and thus may be essential to the secretory process. The monoclonal antibodies will be used to characterize the distribution and membrane organization of the antigens by immunocytochemical and immunochemical techniques and to immunopurify the polypeptides from parotid membranes. The primary structures of the polypeptides will be deduced by peptide sequencing and cloning from a parotid cDNA library. Three kinds of studies are planned toward the major goal of establishing the function of these granule membrane proteins. First, the identification of polypeptides having related sequences and known functions will be sought by computer- searching of centralized sequence data banks. Second, the effects on secretion will be examined in cultured cells in which full-length cDNAs encoding the proteins have been expressed. Finally, the monoclonal antibodies, as well as polyclonal antibodies generated to immunopurified antigens, will be examined as prospective perturbants of protein discharge using a cell-free analog of parotid secretion granule exocytosis. These studies may define some of the final effectors of salivary protein secretion. As such, they will be of substantial interest to other investigators who are attempting to clarify the integration and transduction of secretory stimuli, and they may lead to new insight concerning the regulation of the macromolecular composition of the fluid phase of the oral cavity.

Agency
National Institute of Health (NIH)
Institute
National Institute of Dental & Craniofacial Research (NIDCR)
Type
Research Project (R01)
Project #
5R01DE009655-02
Application #
3223436
Study Section
Oral Biology and Medicine Subcommittee 1 (OBM)
Project Start
1991-01-01
Project End
1995-12-31
Budget Start
1992-01-01
Budget End
1992-12-31
Support Year
2
Fiscal Year
1992
Total Cost
Indirect Cost
Name
University of Virginia
Department
Type
Schools of Medicine
DUNS #
001910777
City
Charlottesville
State
VA
Country
United States
Zip Code
22904
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Liao, Haini; Zhang, Jie; Shestopal, Svetlana et al. (2008) Nonredundant function of secretory carrier membrane protein isoforms in dense core vesicle exocytosis. Am J Physiol Cell Physiol 294:C797-809
Liao, Haini; Ellena, Jeff; Liu, Lixia et al. (2007) Secretory carrier membrane protein SCAMP2 and phosphatidylinositol 4,5-bisphosphate interactions in the regulation of dense core vesicle exocytosis. Biochemistry 46:10909-20
Liu, Lixia; Liao, Haini; Castle, Anna et al. (2005) SCAMP2 interacts with Arf6 and phospholipase D1 and links their function to exocytotic fusion pore formation in PC12 cells. Mol Biol Cell 16:4463-72
Castle, Anna; Castle, David (2005) Ubiquitously expressed secretory carrier membrane proteins (SCAMPs) 1-4 mark different pathways and exhibit limited constitutive trafficking to and from the cell surface. J Cell Sci 118:3769-80
Ellena, Jeffrey F; Moulthrop, Jason; Wu, Jing et al. (2004) Membrane position of a basic aromatic peptide that sequesters phosphatidylinositol 4,5 bisphosphate determined by site-directed spin labeling and high-resolution NMR. Biophys J 87:3221-33
Liu, Lixia; Guo, Zhenheng; Tieu, Quyen et al. (2002) Role of secretory carrier membrane protein SCAMP2 in granule exocytosis. Mol Biol Cell 13:4266-78
Guo, Zhenheng; Liu, Lixia; Cafiso, David et al. (2002) Perturbation of a very late step of regulated exocytosis by a secretory carrier membrane protein (SCAMP2)-derived peptide. J Biol Chem 277:35357-63
Castle, J David; Guo, Zhenheng; Liu, Lixia (2002) Function of the t-SNARE SNAP-23 and secretory carrier membrane proteins (SCAMPs) in exocytosis in mast cells. Mol Immunol 38:1337-40
Hubbard, C; Singleton, D; Rauch, M et al. (2000) The secretory carrier membrane protein family: structure and membrane topology. Mol Biol Cell 11:2933-47

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