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PMID: 2304462 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Promoter and 11-kilobase upstream enhancer elements responsible for hepatoma cell-specific expression of the rat ornithine transcarbamylase gene.

Molecular and cellular biology ·Vol. 10 ·No. 3 ·1990-03-00 ·Pages 1180-91

Murakami T, Nishiyori A, Takiguchi M, Mori M

Abstract

The gene for ornithine transcarbamylase (OTC; EC 2.1.3.3), a urea cycle enzyme, is expressed almost exclusively in the liver and small intestine. To identify DNA elements regulating transcription of the OTC gene in the liver, transient expression analysis was carried out by using hepatoma (HepG2) and nonhepatic (CHO) cell lines. The 1.3-kilobase 5'-flanking region of the rat OTC gene directed expression of the fused chloramphenicol acetyltransferase gene in HepG2 cells much more efficiently than in CHO cells. Analysis of deletion mutants of the 5'-flanking region in HepG2 cells revealed that there are at least one negative and two positive regulatory elements within the about 220-base-pair immediate 5'-flanking region. DNase I footprint analysis showed the presence of factors binding to these regulatory elements in nuclear extracts of rat liver and brain, and footprint profiles at the two positive elements exhibited liver-specific features. Transient expression analysis also revealed the existence of an enhancer region located 11 kilobases upstream of the transcription start site. The OTC enhancer was able to activate both its own and heterologous promoters in HepG2 but not in CHO cells. The enhancer was delimited to an about 230-base-pair region, and footprint analysis of this region revealed four protected areas. Footprint profiles at two of the four areas exhibited liver-specific features, and gel shift competition analysis showed that a factor(s) binding to the two liver-specific sites is related to C/EBP. These results suggest that both liver-specific promoter and enhancer elements regulate expression of the OTC gene through interaction with liver-specific factors binding to these elements.

MeSH Terms
Animals Base Sequence Cell Line Cricetinae DNA-Binding Proteins/physiology Enhancer Elements, Genetic Gene Expression Regulation, Enzymologic Liver Neoplasms, Experimental/genetics Molecular Sequence Data Ornithine Carbamoyltransferase/genetics Promoter Regions, Genetic Rats Regulatory Sequences, Nucleic Acid Restriction Mapping Transcription Factors/physiology
Chemicals
DNA-Binding Proteins Transcription Factors Ornithine Carbamoyltransferase
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Murakami T
Institute for Medical Genetics, Kumamoto University Medical School, Japan.
Nishiyori A
Takiguchi M
Mori M
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/mcb.10.3.1180-1191.1990
Published
1990-03-00
Pages
1180-91
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC360991
Subset
IM
External Links
PubMed source DOI: 10.1128/mcb.10.3.1180-1191.1990 PMC Full Text View journal details
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