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PMID: 2513475 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Use of a selectable marker regulated by alpha interferon to obtain mutations in the signaling pathway.

Molecular and cellular biology ·Vol. 9 ·No. 11 ·1989-11-00 ·Pages 4605-12

Pellegrini S, John J, Shearer M, Kerr IM, Stark GR

Abstract

We have selected mutations in genes encoding components of the signaling pathway for alpha interferon (IFN-alpha) by using a specially constructed cell line. The upstream region of the IFN-regulated human gene 6-16 was fused to the Escherichia coli guanine phosphoribosyltransferase (gpt) gene and transfected into hypoxanthine-guanine phosphoribosyltransferase-negative human cells. These cells express gpt only in the presence of IFN-alpha. They grow in medium containing hypoxanthine, aminopterin, and thymidine plus IFN and are killed by 6-thioguanine plus IFN. Two different types of mutants were obtained after treating the cells with mutagens. A recessive mutant, selected in 6-thioguanine plus IFN, was completely resistant to IFN-alpha but responded normally to IFN-gamma and, unexpectedly, partially to IFN-beta. A constitutive mutant, selected in hypoxanthine-aminopterin-thymidine alone, was abnormal in expressing endogenous genes in the absence of IFN. Both types revert infrequently, allowing selection for complementation of the defects by transfection.

MeSH Terms
Blotting, Northern Cell Fusion Cell Line Cloning, Molecular Escherichia coli/enzymology Gene Expression Regulation Humans Hypoxanthine Phosphoribosyltransferase Interferon Type I/metabolism,pharmacology Interferon-gamma/pharmacology Mutation Pentosyltransferases/genetics Signal Transduction Thioguanine/pharmacology Transfection
Chemicals
Interferon Type I Interferon-gamma Pentosyltransferases Hypoxanthine Phosphoribosyltransferase Thioguanine
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Pellegrini S
Imperial Cancer Research Fund, London, United Kingdom.
John J
Shearer M
Kerr I M
Stark G R
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/mcb.9.11.4605-4612.1989
Published
1989-11-00
Pages
4605-12
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC363606
Subset
IM
External Links
PubMed source DOI: 10.1128/mcb.9.11.4605-4612.1989 PMC Full Text View journal details
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