Sequence specific DNA-protein interactions are responsible for the orderly expression of genetic programs. This proposal outlines an approach whose long range goal is a complete description of both the sequence elements that constitute the promoter of the Rous sarcoma virus long terminal repeat (LTR), and of the cellular components that recognize those elements.
The specific aims of the proposal are the following: 1) To facilitate the localization and subsequent characterization of upstream promoter elements, the region between 55 and 230 base pairs upstream from the transcription start point will be analyzed by the """"""""linker scan"""""""" procedure. The functions of the elements uncovered by the linker scan will be assessed by isolating individual elements and studying their activities in defined contexts. Part of this analysis will determine whether isolated elements, in short-term transfection experiments, can affect the ability of an intact promoter to express bacterial cat gene activity. 2) Using hybrid RSV-MMTV promoters conditions will be sought that permit a demonstration of upstream element-dependent transcription in vitro. 3) Information gained from the linker scan analysis will be used to construct probes that should recognize components of the transcription machinery. The probes will be targets in filter binding assays designed to detect those components. 4) Finally, attempts will be made to select genetic variants with altered expression of genes that code for transcriptional regulatory proteins.
| Greuel, B T; Sealy, L; Majors, J E (1990) Transcriptional activity of the Rous sarcoma virus long terminal repeat correlates with binding of a factor to an upstream CCAAT box in vitro. Virology 177:33-43 |
| Selleck, S B; Majors, J (1988) In vivo ""photofootprint"" changes at sequences between the yeast GAL1 upstream activating sequence and ""TATA"" element require activated GAL4 protein but not a functional TATA element. Proc Natl Acad Sci U S A 85:5399-403 |
| Selleck, S B; Majors, J E (1987) In vivo DNA-binding properties of a yeast transcription activator protein. Mol Cell Biol 7:3260-7 |
| Selleck, S B; Majors, J (1987) Photofootprinting in vivo detects transcription-dependent changes in yeast TATA boxes. Nature 325:173-7 |
| Jacks, T; Townsley, K; Varmus, H E et al. (1987) Two efficient ribosomal frameshifting events are required for synthesis of mouse mammary tumor virus gag-related polyproteins. Proc Natl Acad Sci U S A 84:4298-302 |
