Biosynthesis of the polyamines, spermidine and spermine, is regulated through the key enzyme ornithine decarboxylase (ODC) and is invariably enhanced upon activation of animal cells by a variety of stimuli. The level of ODC mRNA is regulated either transcriptionally or post- transcriptionally, depending on the cell type. The signal transduction pathways that control the ODC gene at the transcriptional level are varied, including those involving protein kinase A (PKA), protein kinase C (PKC), protein tyrosine kinases, and steroid hormone receptors. We at present have identified two regions of the ODC promoter that are important for regulated transcription. Region A is centered at -45 bp relative to the site of transcriptional initiation, in a position adjacent to the TATAA box; this element shows strong sequence similarity to the cAMP-regulated element of the somatostatin gene (""""""""CRE""""""""), and binds the cAMP-regulated factor, CREB, when presented as purified recombinant protein or in cell extracts. Mutation of the CRE-like site strongly depresses basal level transcription driven by the ODC promoter. Region B is a 43-bp GC-rich region located between -92 bp and -134 bp. Deletion of this region, leaving Region A intact, has little or no effect on basal activity of the promoter, but interferes with regulation by PKA or phorbol esters. These results suggest that Region B plays an important role in regulation of ODC transcription by the PKA and PKC pathways. A major goal of this proposal is to test this hypothesis by detailed mutational analysis of Region B, correlating the influence of various mutations on regulation of the promoter with changes in protein binding. Proteins identified by this analysis will be isolated and characterized with regard to their regulatory properties, with the objective of understanding how these DNA-binding proteins interact functionally with the signal transduction pathways. The level of ODC mRNA is also upregulated in fibroblasts by the inflammatory mediator IL-1. The mechanism of this regulation will be examined in detail, and if it is at the transcriptional level, cytokine-responsive elements will be identified in the ODC gene. If these elements are different from those already found, proteins binding to them will be characterized. Regulation by other cytokines of interest, such as TNF- alpha and IL-6, will be examined in the same way.

Agency
National Institute of Health (NIH)
Institute
National Institute of Dental & Craniofacial Research (NIDCR)
Type
Specialized Center (P50)
Project #
3P50DE008229-10S2
Application #
6270286
Study Section
Project Start
1997-12-01
Project End
2000-04-30
Budget Start
1997-10-01
Budget End
1998-09-30
Support Year
10
Fiscal Year
1998
Total Cost
Indirect Cost
Name
University of Washington
Department
Type
DUNS #
135646524
City
Seattle
State
WA
Country
United States
Zip Code
98195
Etikala, Anusha; Bruce, Greg; Hudkins, Kelly et al. (2017) LR8 Expression in fibroblasts of healthy and fibrotic human tissues. Biochem Biophys Rep 10:165-171
Valdés De Hoyos, A; Hoz-Rodríguez, L; Arzate, H et al. (2012) Isolation of protein-tyrosine phosphatase-like member-a variant from cementum. J Dent Res 91:203-9
Alvarez-Perez, Marco Antonio; Narayanan, Sampath; Zeichner-David, Margarita et al. (2006) Molecular cloning, expression and immunolocalization of a novel human cementum-derived protein (CP-23). Bone 38:409-19
Bostrom, A; Bharath, H; Saulewicz, A et al. (2005) Cyclosporin a affects signaling events differentially in human gingival fibroblasts. J Dent Res 84:532-6
Grzesik, Wojciech J; Narayanan, A S (2002) Cementum and periodontal wound healing and regeneration. Crit Rev Oral Biol Med 13:474-84
Dale, B A; Kimball, J R; Krisanaprakornkit, S et al. (2001) Localized antimicrobial peptide expression in human gingiva. J Periodontal Res 36:285-94
Saito, M; Iwase, M; Maslan, S et al. (2001) Expression of cementum-derived attachment protein in bovine tooth germ during cementogenesis. Bone 29:242-8
Yokokoji, T; Narayanan, A S (2001) Role of D1 and E cyclins in cell cycle progression of human fibroblasts adhering to cementum attachment protein. J Bone Miner Res 16:1062-7
Komaki, M; Kang, M; Narayanan, A S (2000) Role of MAP kinases p42erk-2/p44erk-1 in cementum-derived attachment-protein-mediated cell attachment. J Dent Res 79:1789-93
Marshall, A J; Niiro, H; Lerner, C G et al. (2000) A novel B lymphocyte-associated adaptor protein, Bam32, regulates antigen receptor signaling downstream of phosphatidylinositol 3-kinase. J Exp Med 191:1319-32

Showing the most recent 10 out of 135 publications