The acute phase response consists of a large number of defensive and adaptive mechanisms that represent a phenomenon of great biologic importance. Our long-term goals have to delineate the mechanisms that mediate induction of the major human acute phase protein C-reactive protein (CRP) in hepatocytes. IL-6 induces transcription of CRP in the human hepatoma cell line Hep 3B through C/EBP and STAT3 transcription factors. We recently found that the re1 protein p50 participates in IL-6induced CRP expression in two unexpected ways: 1) p50 binds to a non-consensus kappaB site overlapping the proximal C/EBP binding site of the CRP promoter to form ternary complexes with C/EBP species, which are required for CRP induction by IL-6. 2) Constitutive binding of p50 to a non-consensus kappaB site adjacent to the STAT3 response element was displaced by STAT3 binding in the presence of IL-6. These findings did not reflect classic NFkappaB activity; 1) CRP expression was inhibited by over-expressed pp65 and 2) p65 was not found in EMSA complexes. These observations give rise to our working hypotheses, that non-classic re1 family members, including p50, affect IL-6 activated transcription factors at two sites in the proximal promoter: at the C/EBP binding site by stabilizing binding of C/EBP proteins and at the STAT3 binding site, by affecting binding of this factor to its cognate site.
Our specific aims, which are designed to delineate the precise molecular mechanisms that mediate transcription factor interactions in CRP induction, are: I. To define the mechanisms by which p50 influences CRP expression at the critical C/EBP binding site centered at -53. We will determine whether binding of p50 to this region affects affinity and stability of C/EBPbeta for binding at this site and will determine whether these factors physically interact. We will determine if p50 stimulates expression of the endogenous CRP gene and will define the domains of p50 and p65 that mediate their activities. II. To define the mechanisms by which p50 influences CRP expression at the STAT3 binding site. II. To define the To define the mechanisms by which p50 influence CRP expression at the STAT3 binding site and whether increasing p50 affinity for its site alters CRP expression. III. To delineate the specific roles played by IL-6 and IL-1beta in regulating expression on the proximal CRP promoter. IV. To investigate the cytokine response elements in the major response region between -256 and -157 of the CRP promoter. These studies have the potential of contributing substantially to an area of great current interest delineation of the mechanisms by which transcription factors, particularly NFkappaB proteins, interact to influence natural promoters and gene expression.

Agency
National Institute of Health (NIH)
Institute
National Institute on Aging (NIA)
Type
Research Project (R01)
Project #
5R01AG002467-23
Application #
6747292
Study Section
Physiological Chemistry Study Section (PC)
Program Officer
Sierra, Felipe
Project Start
1985-02-01
Project End
2005-05-30
Budget Start
2004-06-01
Budget End
2005-05-30
Support Year
23
Fiscal Year
2004
Total Cost
$306,000
Indirect Cost
Name
Case Western Reserve University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
077758407
City
Cleveland
State
OH
Country
United States
Zip Code
44106
Young, Duprane Pedaci; Kushner, Irving; Samols, David (2008) Binding of C/EBPbeta to the C-reactive protein (CRP) promoter in Hep3B cells is associated with transcription of CRP mRNA. J Immunol 181:2420-7
Chakravarty, Kaushik; Hanson, Richard W (2007) Insulin regulation of phosphoenolpyruvate carboxykinase-c gene transcription: the role of sterol regulatory element-binding protein 1c. Nutr Rev 65:S47-56
Cha-Molstad, Hyunjoo; Young, Duprane Pedaci; Kushner, Irving et al. (2007) The interaction of C-Rel with C/EBPbeta enhances C/EBPbeta binding to the C-reactive protein gene promoter. Mol Immunol 44:2933-42
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Jiang, S; Xia, D; Samols, D (2006) Expression of rabbit C-reactive protein in transgenic mice inhibits development of antigen-induced arthritis. Scand J Rheumatol 35:351-5
Kushner, Irving; Rzewnicki, Debra; Samols, David (2006) What does minor elevation of C-reactive protein signify? Am J Med 119:166.e17-28
Mineo, Chieko; Gormley, Andrew K; Yuhanna, Ivan S et al. (2005) FcgammaRIIB mediates C-reactive protein inhibition of endothelial NO synthase. Circ Res 97:1124-31
Chakravarty, Kaushik; Wu, Shwu-Yuan; Chiang, Cheng-Ming et al. (2004) SREBP-1c and Sp1 interact to regulate transcription of the gene for phosphoenolpyruvate carboxykinase (GTP) in the liver. J Biol Chem 279:15385-95
Black, Steven; Agrawal, Alok; Samols, David (2003) The phosphocholine and the polycation-binding sites on rabbit C-reactive protein are structurally and functionally distinct. Mol Immunol 39:1045-54

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