Multiple point mutations in a shuttle vector propagated in human cells: evidence for an error-prone DNA polymerase activity
Mutagenesis was studied at the DNA-sequence level in human fibroblast and lymphoid cells by use of a shuttle vector plasmid, pZ189, containing a suppressor tRNA marker gene. In a series of experiments, 62 plasmids were recovered that had two to six base substitutions in the 160-base-pair marker gene. Approximately 20-30% of the mutant plasmids that were recovered after passing ultraviolet-treated pZ189 through a repair-proficient human fibroblast line contained these multiple mutations. In contrast, passage of ultraviolet-treated pZ189 through an excision-repair-deficient (xeroderma pigmentosum) line yielded only 2% multiple base substitution mutants. Introducing a single-strand nick in otherwise unmodified pZ189 adjacent to the marker, followed by passage through the xeroderma pigmentosum cells, resulted in about 66% multiple base substitution mutants. The multiple mutations were found in a 160-base-pair region containing the marker gene but were rarely found in an adjacent 170-base-pair region. Passing ultraviolet-treated or nicked pZ189 through a repair-proficient human B-cell line also yielded multiple base substitution mutations in 20-33% of the mutant plasmids. An explanation for these multiple mutations is that they were generated by an error-prone polymerase while filling gaps. These mutations share many of the properties displayed by mutations in the immunoglobulin hypervariable regions.
- Research Organization:
- National Cancer Institute, Bethesda, MD
- OSTI ID:
- 6170288
- Journal Information:
- Proc. Natl. Acad. Sci. U.S.A.; (United States), Journal Name: Proc. Natl. Acad. Sci. U.S.A.; (United States) Vol. 84:14; ISSN PNASA
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
Cells
& Tissue Culture
63 RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTANT EFFECTS ON LIVING ORGS. AND BIOL. MAT.
ANIMAL CELLS
BIOLOGICAL EFFECTS
BIOLOGICAL MATERIALS
BIOLOGICAL RADIATION EFFECTS
BIOLOGICAL RECOVERY
BIOLOGICAL REPAIR
BLOOD
BLOOD CELLS
BODY FLUIDS
CELL CONSTITUENTS
CONNECTIVE TISSUE CELLS
DISEASES
DNA POLYMERASES
DNA REPAIR
ELECTROMAGNETIC RADIATION
ENZYME ACTIVITY
ENZYMES
ERRORS
FIBROBLASTS
GENE MUTATIONS
GENETIC EFFECTS
GENETIC RADIATION EFFECTS
LEUKOCYTES
LYMPHOCYTES
MATERIALS
MUTAGENESIS
MUTATIONS
NUCLEIC ACIDS
NUCLEOTIDYLTRANSFERASES
ORGANIC COMPOUNDS
PHOSPHORUS-GROUP TRANSFERASES
PLASMIDS
POLYMERASES
RADIATION EFFECTS
RADIATIONS
RECOVERY
REPAIR
RNA
SKIN DISEASES
SOMATIC CELLS
STRAND BREAKS
TRANSFER RNA
TRANSFERASES
ULTRAVIOLET RADIATION
XERODERMA PIGMENTOSUM