Signal transduction pathways converge ultimately at the level of transcriptional activation to produce specific patterns of gene expression in response to environmental stimuli. The initiation of transcription mediated by these signalling pathways is regulated by the coordinate expression and/ or activation of specific transcription factors that bind to the control regions of genes. Specific insights into the mechanisms underlying transcriptional activation have recently arisen from the studies of the structure and functions of these transcription factors. The CREB/ATF family of transcriptional transactivating proteins has only recently been discovered and appears to provide a link between the regulation of gene expression in response to activators of cellular signalling pathways and the regulation of gene expression by viral transactivating proteins. Understanding the nature and importance of protein/protein interactions in the mechanisms of eukaryotic gene expression is essential to understanding the normal and aberrant regulation of gene transcription. We have recently developed a rapid and sensitive technique which allows the detection of these interactions between mammalian transcription factors. Utilizing 125I-labelled recombinant CREB and ATF-2 peptides to probe western blots of HeLa nuclear extracts, we have identified multiple separate nuclear factors that form specific protein/protein interactions with these leucine zipper-containing transcriptional regulatory proteins. In this application, we propose to utilize this technique to isolate cDNA clones expressing proteins that form specific protein/protein interactions with CREB and/or ATF-2. In addition, we will perform detailed analyses of the DNA-binding and transcriptional characteristics of several CREB/ATF proteins. The assay of protein/protein interactions we have developed will allow a systematic approach to identifying and cloning proteins involved in the control of eukaryotic transcriptional regulation. The identification and characterization of the components of eukaryotic transcription complexes will allow studies that address the molecular mechanisms of normal and aberrant control of cellular gene expression.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
First Independent Research Support & Transition (FIRST) Awards (R29)
Project #
1R29GM045872-01
Application #
3468423
Study Section
Biochemical Endocrinology Study Section (BCE)
Project Start
1991-04-01
Project End
1996-03-31
Budget Start
1991-04-01
Budget End
1992-03-31
Support Year
1
Fiscal Year
1991
Total Cost
Indirect Cost
Name
University of Colorado Denver
Department
Type
Schools of Medicine
DUNS #
065391526
City
Aurora
State
CO
Country
United States
Zip Code
80045
Russell, M; Hoeffler, J P (1996) Signal transduction and the regulation of cell growth. J Investig Dermatol Symp Proc 1:119-22
Jackson, S M; Gutierrez-Hartmann, A; Hoeffler, J P (1995) Upstream stimulatory factor, a basic-helix-loop-helix-zipper protein, regulates the activity of the alpha-glycoprotein hormone subunit gene in pituitary cells. Mol Endocrinol 9:278-91
Ball, N J; Yohn, J J; Morelli, J G et al. (1994) Ras mutations in human melanoma: a marker of malignant progression. J Invest Dermatol 102:285-90
Abdel-Hafiz, H A; Chen, C Y; Marcell, T et al. (1993) Structural determinants outside of the leucine zipper influence the interactions of CREB and ATF-2: interaction of CREB with ATF-2 blocks E1a-ATF-2 complex formation. Oncogene 8:1161-74
Kroll, D J; Sullivan, D M; Gutierrez-Hartmann, A et al. (1993) Modification of DNA topoisomerase II activity via direct interactions with the cyclic adenosine-3',5'-monophosphate response element-binding protein and related transcription factors. Mol Endocrinol 7:305-18
Jackson, S M; Barnhart, K M; Mellon, P L et al. (1993) Helix-loop-helix proteins are present and differentially expressed in different cell lines from the anterior pituitary. Mol Cell Endocrinol 96:167-76
Anderson, R J; Sponsel, H T; Breckon, R et al. (1993) Transforming growth factor-beta 1 regulation of signal transduction in two renal epithelial cell lines. Am J Physiol 265:F584-91
Kroll, D J; Abdel-Malek Abdel-Hafiz, H; Marcell, T et al. (1993) A multifunctional prokaryotic protein expression system: overproduction, affinity purification, and selective detection. DNA Cell Biol 12:441-53
Hoeffler, J P (1992) Structure/function relationships of CREB/ATF proteins. J Invest Dermatol 98:21S-28S
Abdel-Hafiz, H A; Heasley, L E; Kyriakis, J M et al. (1992) Activating transcription factor-2 DNA-binding activity is stimulated by phosphorylation catalyzed by p42 and p54 microtubule-associated protein kinases. Mol Endocrinol 6:2079-89

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