The long term goal of our research is to determine what cellular processes are regulated by the neuronal calcium-binding protein S100alpha and how S100alpha influences the expression of the neuronal cell phenotype. Studies in our laboratory have directly demonstrated that decreased S100alpha expression blocks NGF-induced neurite extension in PC12 cells. In order to determine how changes in S100alpha expression result in altered phenotypes, one must first determine how S100alpha expression is regulated and what cellular process/target proteins are controlled by S100alpha. The studies proposed in this application utilize a molecular approach directly address both of these questions. The first specific aim is to utilize radioimmunoassays, indirect immunofluorescence microscopy, and dot blot hybridization to quantitate the effects of differentiating agents (e.g. NGF, cAMP, and dexamethasone) on S100alpha protein levels, subcellular distribution, binding protein profiles, and steady state mRNA levels. Nuclear runoff transcription, mRNA stability, and pulse-chase assays will be used to determine if transcriptional and/or post-transcriptional mechanisms regulate S100alpha expression. The second specific aim is to complete the isolation and characterization of the rat S100alpha gene locus(s) and to utilize CAT constructs in transient transfection experiments to determine which S100alpha promoter sequences are involved in transcriptional regulation of S100alpha expression in PC12 cells. In the third specific aim, the level of S100alpha protein will be altered by transfection with S100alpha sense and anti-sense expression vectors to determine the step in the NGF differentiation pathway which is controlled by S100alpha. These same clones will be used in specific aim four to determine if tubulin and/or the microtubule associated gamma protein are regulated by S100alpha in living PC12 cells. These studies will provide the first direct information regarding S100alpha expression and function in neuronal cells and represent a major step in ascertaining the role of S100alpha in neurological disease and its therapeutic potential.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
1R01NS030660-01A1
Application #
3417586
Study Section
Neurological Sciences Subcommittee 1 (NLS)
Project Start
1992-12-01
Project End
1996-11-30
Budget Start
1992-12-01
Budget End
1993-11-30
Support Year
1
Fiscal Year
1993
Total Cost
Indirect Cost
Name
University of South Alabama
Department
Type
Schools of Medicine
DUNS #
City
Mobile
State
AL
Country
United States
Zip Code
36688
Zimmer, D B; Chaplin, J; Baldwin, A et al. (2005) S100-mediated signal transduction in the nervous system and neurological diseases. Cell Mol Biol (Noisy-le-grand) 51:201-14
Rustandi, Richard R; Baldisseri, Donna M; Inman, Keith G et al. (2002) Three-dimensional solution structure of the calcium-signaling protein apo-S100A1 as determined by NMR. Biochemistry 41:788-96
Baldisseri, D M; Rustandi, R R; Zhang, Z et al. (1999) 1H, 13C and 15N NMR sequence-specific resonance assignments for rat apo-S100A1(alpha alpha) J Biomol NMR 14:91-2
Landar, A; Rustandi, R R; Weber, D J et al. (1998) S100A1 utilizes different mechanisms for interacting with calcium-dependent and calcium-independent target proteins. Biochemistry 37:17429-38
Zimmer, D B; Cornwall, E H; Reynolds, P D et al. (1998) S100A1 regulates neurite organization, tubulin levels, and proliferation in PC12 cells. J Biol Chem 273:4705-11
Zimmer, D B; Chessher, J; Wilson, G L et al. (1997) S100A1 and S100B expression and target proteins in type I diabetes. Endocrinology 138:5176-83
Landar, A; Hall, T L; Cornwall, E H et al. (1997) The role of cysteine residues in S100B dimerization and regulation of target protein activity. Biochim Biophys Acta 1343:117-29
Landar, A; Caddell, G; Chessher, J et al. (1996) Identification of an S100A1/S100B target protein: phosphoglucomutase. Cell Calcium 20:279-85
Song, W; Zimmer, D B (1996) Expression of the rat S100A1 gene in neurons, glia, and skeletal muscle. Brain Res 721:204-16
Zimmer, D B; Chessher, J; Song, W (1996) Nucleotide homologies in genes encoding members of the S100 protein family. Biochim Biophys Acta 1313:229-38

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