The unusual organization of INK4a/ARF gene locus permits the expression of two overlapping transcripts that are translated in alternate reading frames. Recently we have developed a novel and highly efficient method for the transfer of bacterial artificial chromosomes (BACs) into cells in culture. Combined with the ability to introduce specific modifications in BACs by homologous recombination, this novel gene transfer system provides an opportunity to dissect the role of specific regulatory elements in the context of complex genetic loci, both in vitro and in BAC transgenic mice. Using this gene transfer system and modified alleles of INK4a/ARF, we propose to: 1) investigate the function of a TTTCCCGC E2F binding site in activation of pl9ARF mRNA expression in response to the transcriptional activators E2Fs 1-3, and 2) examine the specific role of E2F-6 in repression of pl9?ARF transcription. These studies will develop a new paradigm for dissecting the role of specific regulatory elements within the context of entire genetic loci in cells in culture, thereby permitting evaluation of specific BAC alleles prior to the generation of BAC transgenic mice.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
National Research Service Awards for Senior Fellows (F33)
Project #
1F33GM064167-01
Application #
6406043
Study Section
Cell Development and Function Integrated Review Group (CDF)
Program Officer
Tompkins, Laurie
Project Start
2002-01-17
Project End
Budget Start
2001-09-15
Budget End
2002-05-14
Support Year
1
Fiscal Year
2001
Total Cost
$34,896
Indirect Cost
Name
University of Vermont & St Agric College
Department
Pathology
Type
Schools of Medicine
DUNS #
066811191
City
Burlington
State
VT
Country
United States
Zip Code
05405