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PMID: 1996095 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Dissection of the mouse N-ras gene upstream regulatory sequences and identification of the promoter and a negative regulatory element.

Molecular and cellular biology ·Vol. 11 ·No. 3 ·1991-03-00 ·Pages 1334-43

Paciucci R, Pellicer A

Abstract

The 5' flanking region of the mouse N-ras gene was investigated to determine the elements governing transcriptional activity of the gene. The promoter did not contain typical TATA or CCAAT boxes, and according to primer extension and RNase protection analyses, transcription started at several sites. These assays also confirmed the short nucleotide distance interposed between the N-ras transcription unit and the previously described upstream unr gene. Chromatin studies performed by digestion of nuclei with DNase I revealed the presence of four hypersensitive sites: a, b, c, and d. Deletion mutagenesis of the 5' flanking region revealed sequences responsible for both promotion and inhibition of transcription. These sequences resided within 230 bp upstream of the transcription initiation site. Hypersensitive site b colocalized with the 76-bp segment with promoter activity. The negative regulatory element at position -180 colocalized with hypersensitive site a, was active on the N-ras promoter in stable as well as transient assays, and down-regulated the heterologous herpes simplex virus thymidine kinase promoter. Footprint analysis and in vivo transfection-competition experiments indicated that a trans-acting factor is responsible for the negative effect on transcription. The interaction between the cis-acting negative regulatory element and the promoter region may play a role in the tissue- and developmental-stage-specific patterns of expression of the N-ras gene.

Related Genes
N-ras
MeSH Terms
Animals Base Sequence DNA Mutational Analysis DNA-Binding Proteins/physiology Mice Molecular Sequence Data Oligonucleotides/chemistry Promoter Regions, Genetic Proto-Oncogene Proteins p21(ras)/genetics Proto-Oncogenes Regulatory Sequences, Nucleic Acid Structure-Activity Relationship Transcription Factors/physiology Transcription, Genetic
Chemicals
DNA-Binding Proteins Oligonucleotides Transcription Factors Proto-Oncogene Proteins p21(ras)
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Paciucci R
Department of Pathology, New York University Medical Center, New York 10016.
Pellicer A
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/mcb.11.3.1334-1343.1991
Published
1991-03-00
Pages
1334-43
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC369404
Subset
IM
Grants
NCI NIH HHS · CA 36327 · United States
NCI NIH HHS · CA 50434 · United States
External Links
PubMed source DOI: 10.1128/mcb.11.3.1334-1343.1991 PMC Full Text View journal details
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