Document Detail


The human U1 snRNA promoter correctly initiates transcription in vitro and is activated by PSE1.
MedLine Citation:
PMID:  2125285     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
A DNA-dependent in vitro transcription system for the human U1 small nuclear RNA (snRNA) promoter has been developed. This in vitro transcription system uses extracts of tissue culture cells to drive transcription of an RNA polymerase II-transcribed snRNA gene. A U1 promoter (-393 to +192) template was constructed in which the sequences from +10 to +171 were replaced with a 179-bp sequence from a G-less cassette. This DNA template thus retained all of the known U1 promoter elements, including the U1 3'-end box (positions +175 to +191), which is responsible for snRNA 3'-end formation. HeLa cell nuclear extracts were shown to drive specific transcription of this promoter by RNA polymerase II. This transcription system has many of the properties observed for wild-type snRNA promoters in vivo. Transcription was shown to initiate at +1 (and -2) relative to the U1 promoter and to efficiently (greater than 90%) form a 3' end corresponding to the 3' end found in the primary transcript of U1 in vivo. The transcription signal is responsive to either deletion or replacement of the U1 distal sequence (enhancer-like) and proximal sequence (TATA-like) elements, as well as the 3'-end box. Additionally, the signal was shown by depletion/repletion experiments to be responsive to a protein called PSE1 (related to Ku), which has recently been shown to specifically bind sequences in the U1 promoter. This in vitro snRNA transcription system should facilitate the biochemical analysis of the human U1 snRNA promoter and lead to a better understanding of the differences between snRNA and mRNA promoters.
Authors:
S I Gunderson; M W Knuth; R R Burgess
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Publication Detail:
Type:  Journal Article    
Journal Detail:
Title:  Genes & development     Volume:  4     ISSN:  0890-9369     ISO Abbreviation:  Genes Dev.     Publication Date:  1990 Dec 
Date Detail:
Created Date:  1991-02-19     Completed Date:  1991-02-19     Revised Date:  2008-11-21    
Medline Journal Info:
Nlm Unique ID:  8711660     Medline TA:  Genes Dev     Country:  UNITED STATES    
Other Details:
Languages:  eng     Pagination:  2048-60     Citation Subset:  IM    
Affiliation:
McArdle Laboratory for Cancer Research, University of Wisconsin, Madison 53706.
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MeSH Terms
Descriptor/Qualifier:
Base Sequence
Cloning, Molecular
DNA-Binding Proteins / metabolism*
Guanosine Triphosphate / analogs & derivatives,  metabolism
Hela Cells
Humans
Molecular Sequence Data
Mutation
Promoter Regions, Genetic*
Proteins*
RNA Polymerase II / metabolism
RNA, Small Nuclear / genetics*
Ribonuclease T1 / metabolism
Templates, Genetic
Transcription, Genetic*
Chemical
Reg. No./Substance:
0/DNA-Binding Proteins; 0/PSE1 protein, human; 0/Proteins; 0/RNA, Small Nuclear; 61556-45-4/3'-O-methylguanosine triphosphate; 86-01-1/Guanosine Triphosphate; EC 2.7.7.-/RNA Polymerase II; EC 3.1.27.3/Ribonuclease T1

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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