The necessary intricacy of gene expression in higher metazoans requires a myriad of positive and negative regulators for temporal and cell-specific control of transcription. Despite this complexity, the core promoter TATA- element binding protein, TBP, is a ubiquitous, highly-conserved central component of the general transcription machinery of all eukaryotic promoters. Recently, a Drosophila protein with striking homology to dTBP, dTBP-related factor (dTRF), was identified and found, surprisingly, to be highly tissue-specific. This raise the intriguing possibility that some transcriptional regulators may utilize core promoter complexes other than those containing TBP and TFIID to direct cell-type specific expression of genes. This research plan will investigate the potential role dTRF plays in tissue-specific transcriptional control. Specific objectives are: (1) Identification of a functional dTRF complex and its associated factors. Immunopurification and Far Western blot assays are proposed to isolate potential dTRF-associated factors. (2) Identification of dTRF-regulated target genes. A random site-selection is proposed to identify high affinity of dTRF recognition elements later used to determine functionality and to identify potential Drosophila genes under transcriptional control by dTRF. (3) Functional relevance of dTRF:promoter context through in vivo and in vitro analysis. Cell culture models of dTRF-mediated gene control will be tested by assay of reporter constructs with putative dTRF targets. In addition, biochemical characterization of dTRF-dependent transcription through cell-free reconstitution is proposed. The recapitulation of regulated transcription of a bona-fide dtrf-target gene could provide novel insights into mechanisms employed by metazoans for tissue-specific gene expression.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Postdoctoral Individual National Research Service Award (F32)
Project #
5F32GM019325-03
Application #
6179211
Study Section
Biological Sciences 2 (BIOL)
Program Officer
Tompkins, Laurie
Project Start
1998-06-01
Project End
Budget Start
2000-06-01
Budget End
2001-05-31
Support Year
3
Fiscal Year
2000
Total Cost
$37,516
Indirect Cost
Name
University of California Berkeley
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
094878337
City
Berkeley
State
CA
Country
United States
Zip Code
94704