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Mol Cell Biol, March 1998, p. 1190-1200, Vol. 18, No. 3
Department of Biochemistry and Molecular
Biology, The Albany Medical College, Albany, New York
12208-3479,1 and
Department of
Radiotherapy, Loyola University Medical Center, Maywood, Illinois
601532
Received 6 October 1997/Returned for modification 11 November
1997/Accepted 26 November 1997
Genetic instability in the Saccharomyces cerevisiae
rad9 mutant correlates with failure to arrest the cell cycle in
response to DNA damage. We quantitated the DNA damage-associated
stimulation of directed translocations in RAD9+
and rad9 mutants. Directed translocations were generated by
selecting for His+ prototrophs that result from homologous,
mitotic recombination between two truncated his3 genes,
GAL1::his3-
0270-7306/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.
The Saccharomyces cerevisiae RAD9
Checkpoint Reduces the DNA Damage-Associated Stimulation of
Directed Translocations
5' and
trp1::his3-
3'::HOcs. Compared to
RAD9+ strains, the rad9 mutant
exhibits a 5-fold higher rate of spontaneous, mitotic recombination and
a greater than 10-fold increase in the number of UV- and
X-ray-stimulated His+ recombinants that contain
translocations. The higher level of recombination in rad9
mutants correlated with the appearance of nonreciprocal translocations
and additional karyotypic changes, indicating that genomic instability
also occurred among non-his3 sequences. Both enhanced
spontaneous recombination and DNA damage-associated recombination are
dependent on RAD1, a gene involved in DNA excision repair.
The hyperrecombinational phenotype of the rad9 mutant was
correlated with a deficiency in cell cycle arrest at the
G2-M checkpoint by demonstrating that if rad9
mutants were arrested in G2 before irradiation, the numbers
both of UV- and
-ray-stimulated recombinants were reduced. The
importance of G2 arrest in DNA damage-induced sister
chromatid exchange (SCE) was evident by a 10-fold reduction in HO
endonuclease-induced SCE and no detectable X-ray stimulation of SCE in
a rad9 mutant. We suggest that one mechanism by which the
RAD9-mediated G2-M checkpoint may reduce the
frequency of DNA damage-induced translocations is by channeling the
repair of double-strand breaks into SCE.
*
Corresponding author. Present address: Department of
Biochemistry and Molecular Biology, The Albany Medical College, 47 New Scotland Ave., Albany, NY 12208-3479. Phone: (518) 262-6651. Fax: (518)
262-5689. E-mail: mfasullo{at}ccgateway.amc.edu.
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