Location of S1 Nuclease-Cleavage Sites on Circular, Superhelical DNAs between Polyoma Virus and Simian Virus 40

In addition to the highly conserved AATAAA sequence, there is a requirement for specific sequences downstream of polyadenylic acid [poly(A)] cleavage sites to generate correct mRNA 3' termini. Previous experiments demonstrated that 35 nucleotides downstream of the E2A poly(A) site were sufficient but 20 nucleotides were not. The construction and assay of bidirectional deletion mutants in the adenovirus E2A poly(A) site indicates that there may be redundant multiple sequence elements that affect poly(A) site usage. Sequences between the poly(A) site and 31 nucleotides downstream were not essential for efficient cleavage. Further deletion downstream (3' to +31) abolished efficient cleavage in certain constructions but not all. Between +20 and +38 the sequence T(A/G)TTTTT was duplicated. Function was retained when one copy of the sequence was present, suggesting that this sequence represents an essential element. There may also be additional sequences distal to +43 that can function. To establish common features of poly(A) sites, we also analyzed the early simian virus 40 (SV40) poly(A) site for essential sequences. An SV40 poly(A) site deletion that retained 18 nucleotides downstream of the cleavage site was fully functional while one that retained 5 nucleotides downstream was not, thus defining sequences required for cleavage. Comparison of the SV40 sequences with those from E2A did not reveal significant homologies. Nevertheless, normal cleavage and polyadenylation could be restored at the early SV40 poly(A) site by the addition of downstream sequences from the adenovirus E2A poly(A) site to the SV40 +5 mutant. The same sequences that were required in the E2A site for efficient cleavage also restored activity to the SV40 poly(A) site.

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A specific DNA sequence controls termination of transcription in the gastrin gene

Molecular and Cellular Biology ◽

10.1128/mcb.6.4.1032-1043.1986 ◽

1986 ◽

Vol 6 (4) ◽

pp. 1032-1043

Author(s):

K Sato ◽

R Ito ◽

K H Baek ◽

K Agarwal

Keyword(s):

Signal Sequence ◽

Regulatory Element ◽

Shuttle Vector ◽

Simian Virus 40 ◽

Simian Virus ◽

Regulation Of Transcription ◽

Dependent Manner ◽

Transcriptional Terminator ◽

S1 Nuclease ◽

Nuclease Mapping

We located and characterized a downstream transcriptional regulatory element in the human gastrin gene by transferring the gastrin gene 3' fragment, from which the polyadenylation signal sequence was deleted, into the shuttle vector pSCAT10 at a site located immediately downstream from the chloramphenicol acetyltransferase (CAT) gene and upstream from the simian virus 40 polyadenylation region. Study of CAT RNA derived from the hybrid plasmids, indicated regulation of transcription on the gastrin gene fragment. Analysis of deletion mutants generated from the 5' region of the fragment by CAT assay and by S1 nuclease mapping of mRNAs indicated the possible involvement of an oligothymidylate-rich sequence in transcription regulation. Mapping of gastrin gene RNA 3' ends to the 5' side proximal to the oligothymidylate-rich sequence clearly demonstrated that this sequence is a transcriptional terminator element. This unique sequence, interspersed with one or two adenines, which also functions in an orientation-dependent manner, is located 192 nucleotides downstream from the gastrin gene polyadenylation site, and serves as a transcriptional termination signal.

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Lack of Sequence Homology Between the 70S RNA of Various RNA Tumor Viruses and the DNA of Simian Virus 40 or Polyoma Virus

Journal of Virology ◽

10.1128/jvi.11.6.1027-1029.1973 ◽

1973 ◽

Vol 11 (6) ◽

pp. 1027-1029

Author(s):

Robert E. Gallagher ◽

Arthur S. Levine ◽

David H. Gillespie ◽

Robert C. Gallo

Keyword(s):

Sequence Homology ◽

Simian Virus 40 ◽

Simian Virus ◽

Polyoma Virus ◽

Tumor Viruses ◽

Rna Tumor Viruses

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Hormonally mediated negative regulation of human pro-opiomelanocortin gene expression after transfection into mouse L cells.

Molecular and Cellular Biology ◽

10.1128/mcb.5.9.2443 ◽

1985 ◽

Vol 5 (9) ◽

pp. 2443-2453 ◽

Cited By ~ 30

Author(s):

A Israel ◽

S N Cohen

Keyword(s):

Simian Virus 40 ◽

Simian Virus ◽

Negative Regulation ◽

Coding Region ◽

Transient Expression Assay ◽

S1 Nuclease ◽

Protein Coding ◽

Pomc Gene ◽

Simplex Virus ◽

Nuclease Mapping

We report results indicating that expression and hormonally controlled negative regulation of the human pro-opiomelanocortin (POMC) gene in mouse fibroblasts can be accomplished by the placement nearby of a simian virus 40 enhancer sequence. Expression resulting from correctly initiated transcription required the enhancer in cis both in cells stably transfected with the POMC gene and in a transient expression assay with constructs that fused that POMC promoter region to the protein-coding region of the herpes simplex virus thymidine kinase (TK) gene. Negative regulation of POMC transcription by glucocorticoids was demonstrated in transiently infected cells by assaying for TK activity encoded by the POMC-TK fusion constructs and by quantitative S1 nuclease mapping. The sequences responsible for such regulation were shown to be contained within a DNA segment that extends 670 base pairs upstream from the cap site for POMC mRNA.

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