# MECHANISM OF MUTATION INDUCED IN MAMMALIAN GENE

> **NIH NIH R01** · COLUMBIA UNIV NEW YORK MORNINGSIDE · 1992 · $187,868

## Abstract

The proposal describes approaches to define the spectrum of
mutations induced two model chemical carcinogens in an endogenous
mammalian gene at the DNA sequence level.  The carcinogens we used
were N-acetoxy-N-2-acetylaminofluorene (AAAF) and (plus/minus)-r-
7,t-8-dihydroxy-t-9, lO-epoxy-7,8,9,10-tetrahydrobenzo(a)pyrene
(BPDE).  The target for mutation was the dominant autosomal
dihydrofolate reductase (dhfr) gene of Chinese hamster ovary cells.
DHFR- mutants are triple auxotrophs for glycine, thymidine, and
hypoxanthine and were selected starting with a hemizygote cell line
(dhfr+/delta).  Characterization of the induced mutants by Southern
blot analysis showed that in the case of AAAF, a significant number
(28%) carried gene rearrangements (deletion, translocation, large
insertion, or inversion).  In the case of BPDE, the dhfr- mutants
virtually all carried small lesions (base substitution, + or -1
frameshift mutation).  We also examined the expression phenotype
of the induced mutants by Northern analysis and RNA heteroduplex
mapping.  About 50% of the putative point mutants showed markedly
reduced steady-state levels of poly(A)+ dhfr mRNA.  By cloning and
sequencing, we determined that two mutants displaying a low mRNA
phenotype carried chain termination codons at different positions
in the dhfr coding sequence.  Our proposal describes plans for the
further study of these mutants.  We describe methods to facilitate
the DNA sequence analysis of the putative point mutants.  We intend
to isolate dhfr- mutants induced with a related aromatic amine, N-
hydroxy-AF in order to determine if the mutational spectra of AAAF
is associated with the conformational changes in DNA structure
induced by the dG-adduct.  In collaboration, we will determine if
chromosomal location influences the frequency and spectra of
induced mutations.  Finally, we will attempt to suppress the
nonsense mutations by transfection of Su+ tRNA and selection for
a DHFR+ phenotype.  We will then evaluate the dhfr mRNA levels in
these transfected cells.

## Key facts

- **NIH application ID:** 3178659
- **Project number:** 5R01CA039547-08
- **Recipient organization:** COLUMBIA UNIV NEW YORK MORNINGSIDE
- **SAM.gov UEI:** F4N1QNPB95M4
- **Principal Investigator:** DEZIDER GRUNBERGER
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 1992
- **Award amount:** $187,868
- **Award type:** 5
- **Project period:** 1985-04-01 → 1996-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/3178659

## Citation

> US National Institutes of Health, RePORTER application 3178659, MECHANISM OF MUTATION INDUCED IN MAMMALIAN GENE (5R01CA039547-08). Retrieved via MORVS 2026-10-25 from https://api.morvs.ai/grant/nih/3178659. Licensed CC0.

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