There is good evidence from our laboratory indicating that catecholamines through cAMP (adenosine 3 feet, 5 feet-monophosphate) mediate the induction of the lactate dehydrogenase M subunit in rat 06 glioma cells at the transcriptional level. However, the mechanism whereby cAMP initiates and controls the transcriptional events is as yet unknown, but based on its established effects may involve a mechanism of activation of nuclear cAMP-dependent protein kinase (through nuclear translocation of cytoplasmic cAMP-dependent protein kinase), phosphorylative modification of nucleosomal proteins and other nuclear proteins resulting in a structurally and functionally altered chromatin. The proposed studies are aimed at defining (a) the mechanism of nuclear translocation of cytoplasmic cAMP-dependent protein kinase by biochemical and immunochemical methods; (b) to identify nuclear protein kinase (and phosphatase) substrates and the exact phosphorylation sites; (c) to elucidate the kinetics of phosphorylation/ (dephosphorylation after isoproterenol stimulation; and (d) to functionally correlate these phosphorylation/dephosphorylation events with induction of nuclear lactate dehydrogenase mRNA synthesis after isoproterenol stimulation.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM023895-06
Application #
3271917
Study Section
Pathobiochemistry Study Section (PBC)
Project Start
1978-01-01
Project End
1986-08-31
Budget Start
1985-09-01
Budget End
1986-08-31
Support Year
6
Fiscal Year
1985
Total Cost
Indirect Cost
Name
Northwestern University at Chicago
Department
Type
School of Medicine & Dentistry
DUNS #
005436803
City
Chicago
State
IL
Country
United States
Zip Code
60611
Short, M L; Manohar, C F; Furtado, M R et al. (1991) Nucleotide and derived amino-acid sequences of the CRE-binding proteins from rat C6 glioma and HeLa cells. Nucleic Acids Res 19:4290
Mednieks, M I; Jungmann, R A; Fischler, C et al. (1989) Immunogold localization of the type II regulatory subunit of cyclic AMP-dependent protein kinase. Monoclonal antibody characterization and RII distribution in rat parotid cells. J Histochem Cytochem 37:339-46
Kwast-Welfeld, J; Soong, C J; Short, M L et al. (1989) Identification of rat ovarian nuclear factors that interact with the cAMP-inducible lactate dehydrogenase A subunit promoter. J Biol Chem 264:6941-7
Squinto, S P; Jungmann, R A (1989) Modulation of nuclear cyclic AMP-dependent protein kinase in dibutyryl cyclic AMP-treated rat H4IIE hepatoma cells. Biochem J 260:673-82
Hand, A R; Jungmann, R A (1989) Localization of cellular regulatory proteins using postembedding immunogold labeling. Am J Anat 185:183-96
Jungmann, R A; Kuettel, M R; Squinto, S P et al. (1988) Using immunocolloidal gold electron microscopy to investigate cAMP-dependent protein kinase cellular compartmentalization. Methods Enzymol 159:225-35
Jungmann, R A; Constantinou, A I; Squinto, S P et al. (1986) Regulation of lactate dehydrogenase gene expression by cAMP-dependent protein kinase subunits. Ann N Y Acad Sci 478:147-61
Constantinou, A I; Squinto, S P; Jungmann, R A (1985) The phosphoform of the regulatory subunit RII of cyclic AMP-dependent protein kinase possesses intrinsic topoisomerase activity. Cell 42:429-37
Squinto, S P; Kelley-Geraghty, D C; Kuettel, M R et al. (1985) Ultrastructural localization of cAMP-dependent protein kinase subunits in regenerating rat hepatocytes using immunogold electron microscopy. J Cyclic Nucleotide Protein Phosphor Res 10:65-73
Kuettel, M R; Squinto, S P; Kwast-Welfeld, J et al. (1985) Localization of nuclear subunits of cyclic AMP-dependent protein kinase by the immunocolloidal gold method. J Cell Biol 101:965-75