Enhancers can be located thousands of base pairs from a gene promoter and still stimulate transcription. Although there is no definitive data on how enhancers act over such long distances, enhancers are widely believed to serve as the binding sites for transcription factors which migrate to gene promoters via a looping out of the intervening DNA. In contrast to this conventional view, we have obtained evidence suggesting that the c-fos and polyoma enhancers act by inducing a greater accessibility of the DNA template as detected by increased DNase I sensitivity. A major goal of this proposal is to test the model that enhancers act at a distance by increasing the genetic accessibility of chromatin. To address this objective, enhancers or enhancer deletions will be inserted into the hypoxanthine phosphoribosyl transferase (HPRT) gene of male human diploid cells by site-directed homologous recombination (gene targeting). Nuclei from recombinant or wild type cells resulting from the DNA inserts. Relative rates of DNase I sensitivity will be determined by a novel PCR assay that reliably measures DNA accessibility at any genetic loci in chromatin. These experiments will determine definitively whether enhancers can elevate DNase I sensitivity at a locus characterized by low native DNase I sensitivity.***//

Agency
National Science Foundation (NSF)
Institute
Division of Molecular and Cellular Biosciences (MCB)
Application #
9018701
Program Officer
DeLill Nasser
Project Start
Project End
Budget Start
1991-06-01
Budget End
1993-05-31
Support Year
Fiscal Year
1990
Total Cost
$194,907
Indirect Cost
Name
University of Michigan Ann Arbor
Department
Type
DUNS #
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109