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Deletion and mutation studies of the human 7SK gene transfected into HeLa cells have identified three functional regions of the promoter corresponding to the TATA box at -25, the proximal sequence element (PSE) between -49 and -65 and the distal sequence element (DSE) between -243 and -210. These elements show sequence homology to equivalent regions in other snRNA genes and are functionally analogous. Unlike the DSEs of many snRNA genes however, the 7SK DSE does not contain a consensus binding site for the transcription factor Oct-1 but rather, contains two non-consensus Oct-1 binding sites that can function independently of one another to enhance transcription. Unusually, the 7SK PSE can retain function even after extensive mutation and removal of the conserved TGACC of the PSE has little effect in the context of the whole promoter. However, the same mutation abolishes transcription in the absence of the DSE suggesting that protein/protein interactions between DSE and PSE binding factors can compensate for a mutant PSE. Mutation of the 7SK TATA box allows snRNA type transcription by RNA polymerase II to occur and this is enhanced by the DSE, indicating that both the DSE and the PSE can also function with pol II. In addition, mutation of the TATA box does not abolish pol III dependent transcription, suggesting that other sequence elements may also play a role in the determination of polymerase specificity. Although the human 7SK gene is transcribed efficiently in Xenopus oocytes, analysis of the 7SK wild-type gene and mutants in Xenopus oocytes gives significantly different results from the analysis in HeLa cells indicating that the recognition of functional elements is not the same in the two systems.

Original publication

DOI

10.1006/jmbi.1995.0582

Type

Journal article

Journal

J Mol Biol

Publication Date

10/11/1995

Volume

253

Pages

677 - 690

Keywords

Amino Acid Sequence, Animals, Base Sequence, DNA-Binding Proteins, Enhancer Elements, Genetic, HeLa Cells, Host Cell Factor C1, Humans, Macromolecular Substances, Molecular Sequence Data, Mutagenesis, Octamer Transcription Factor-1, Oocytes, Promoter Regions, Genetic, RNA Polymerase II, RNA Polymerase III, RNA, Small Nuclear, Recombinant Fusion Proteins, Sequence Deletion, TATA Box, Transcription Factor TFIID, Transcription Factors, Transcription, Genetic, Transfection, Xenopus Proteins, Xenopus laevis