Ultraviolet-endonuclease activity in cell extracts of Saccharomyces cerevisiae mutants defective in excision of pyrimidine dimers

Bekker, M L; Kaboev, O. K.; Akhmedov, Alexander; Luchkina, L A

doi:10.1128/jb.142.1.322-324.1980

Cited by 19 publications

(11 citation statements)

References 14 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Further studies on the dimer-excising activity in cell-free systems will be reported elsewhere (R. J. Reynolds and E. C. Friedberg, manuscript in preparation). These conclusions are not in agreement with the recent report by Bekker et al (2) who claim to have demonstrated normal levels of "UV endonuclease" activity in extracts of a number of rad mutants of S. cerevisiae. Our results with mutants at the RAD7 and RAD14 loci are not definitive, nor have we yet examined mutants at the RADIO, RAD16, or MMS19 loci.…”

Section: Discussioncontrasting

confidence: 96%

Molecular Mechanisms of Pyrimidine Dimer Excision in Saccharomyces cerevisiae : Incision of Ultraviolet-Irradiated Deoxyribonucleic Acid In Vivo

1981

View full text Add to dashboard Cite

A group of genetically related ultraviolet (UV)-sensitive mutants of Saccharomyces cerevisiae has been examined in terms of their survival after exposure to UV radiation, their ability to carry out excision repair of pyrimidine dimers as measured by the loss of sites (pyrimidine dimers) sensitive to a dimer-specific enzyme probe, and in terms of their ability to effect incision of their deoxyribonucleic acid (DNA) during post-UV incubation in vivo (as measured by the detection of single-strand breaks in nuclear DNA). In addition to a haploid RAD+ strain (S288C), 11 different mutants representing six RAD loci (RADI, RAD2, RAD3, RAD4, RAD14, and RAD18) were examined. Quantitative analysis of excision repair capacity, as determined by the loss of sites in DNA sensitive to an enzyme preparation from M. luteus which is specific for pyrimidine dimers, revealed a profound defect in this parameter in all but three of the strains examined. The radl4-1 mutant showed reduced but significant residual capacity to remove enzyme-sensitive sites as did the rad2-4 mutant. The latter was the only one of three different rad2 alleles examined which was leaky in this respect. The UV-sensitive strain carrying the mutant allele radl8-1 exhibited normal loss of enzyme-sensitive sites consistent with its ament to the RAD6 rather than the RAD3 epistatic group. All strains having mutant alleles of the RADI, RAD2, RAD3, RAD4, and RAD14 loci showed no detectable incubation-dependent strand breaks in nuclear DNA after exposure to UV radiation. These experiments suggest that the RADI, RAD2, RAD3, RAD4 (and probably RAD14) genes are aJl required for the incision of UV-irradiated DNA during pyrimidine dimer excision in vivo.

show abstract

Section: Discussioncontrasting

confidence: 96%

Molecular Mechanisms of Pyrimidine Dimer Excision in Saccharomyces cerevisiae : Incision of Ultraviolet-Irradiated Deoxyribonucleic Acid In Vivo

1981

View full text Add to dashboard Cite

show abstract

“…The levels of these activities were the same for all the strains studied and corresponded well to the in vivo excision capacity of wild-type cells. Our previous data [5] were confirmed in the special sets of experiments.…”

Section: Published By Elsevier Science Publishers B Vsupporting

confidence: 78%

“…However this suggestion does not seem to be valid in the light of data that were obtained with a double mutant, which carried both the rad mutation and the cdc9 mutation, that is also defective in DNA ligase [4]. Despite the absence of in vivo incision we showed that cell-free extracts of RAD3 group mutants contained a nuclease activity specific for UV-irradiated Co1 E 1 DNA [5]. The level of this activity did not differ from that in wild-type extracts.…”

Section: Introductioncontrasting

confidence: 66%

“…On the other hand, they reported that the extracts of both wildtype yeast and RAD3 group mutants catalyzed the excision of PD from UV-irradiated DNA, that had been pretreated with Micrococcus luteus UVendonuclease activity [6]. They noted in [l] that their data disagree with ours [5]. This controversy prompted us to assay both qualitatively and quantitatively in vitro the PD excision activity in extracts of wild-type and rad mutant cells using UVirradiated DNA that had not been pretreated with exogenous UV-endonuclease.…”

Section: Published By Elsevier Science Publishers B Vmentioning

confidence: 77%

“…UV-endonuclease activity was determined by agarose gel electrophoresis, using 14C-labeled Co1 El DNA as in [5]. UV-irradiated DNA as a substrate for this study was prepared by irradiation of Co1 El DNA at 254 nm at 60 J/m*.…”

Section: Methodsmentioning

confidence: 99%

See 2 more Smart Citations

Quantitative characterization of pyrimidine dimer excision from UV‐irradiated DNA (excision capacity) by cell‐free extracts of the yeast Saccharomyces cerevisiae

et al. 1984

View full text Add to dashboard Cite

Cell-free extracts from wild-type yeast @AD+) and from rad mutants belonging to the R4D3 epistatic group (radl-1, rad2-1, rad3-1, rad4-1) contain activities catalyzing the excision of pyrimidine dimers (PD) from purified ultraviolet-irradiated DNA which was not pre-treated with exogenous UV-endonuclease. The level of these activities in cell-free extracts from rad mutants did not differ from that in wild-type extract and was close to the in vivo excision capacity of the latter calculated from the LDH (about lo4 PD per haploid genome).

show abstract

Repair of UV-irradiated plasmid DNA in a Saccharomyces cerevisiae rad3 mutant deficient in excision-repair of pyrimidine dimers

Domiński

Jachmczyk

1984

Molec. Gen. Genet.

View full text Add to dashboard Cite

The repair of UV-irradiated DNA of plasmid pBB29 was studied in an incision-defective rad3-2 strain of Saccharomyces cerevisiae and in a uvrA6 strain of Escherichia coli by the measurement of cell transformation. Plasmid pBB29 used in these experiments contained as markers the DNA of nuclear yeast gene LEU-2 and DNA of the bacterial plasmid pBR327 with resistance to Tet and Amp enabling simultaneous screening of transformant cells in both microorganisms. We found that the yeast rad3-2 mutant, deficient in incision of UV-induced pyrimidine dimers in nuclear DNA, was fully capable of repairing such lesions in plasmid DNA. The repair efficiency was comparable to that of the wild-type cells. The E. coli uvrA6 mutant, deficient in a specific nuclease for pyrimidine dimer excision from chromosomal DNA, was unable to repair UV-damaged plasmid DNA. The difference in repair capacity between the uvrA6 mutant strain and the wild-type strain was of several thousand-fold. It seems that the rad3 mutation, which confers deficiency in the DNA excision-repair system in yeast, is limited only to the nuclear DNA.

show abstract

Ultraviolet-endonuclease activity in cell extracts of Saccharomyces cerevisiae mutants defective in excision of pyrimidine dimers

Abstract: Cell-free extracts of ultraviolet-sensitive mutants of Saccharomyces cerevisiae defective in excision of pyrimidine dimers, rad1, rad2, rad3, rad4, rad10, and rad16, as well as the extracts of the wild-type strain RAD+, display ultraviolet-endonuclease activity.

Cited by 19 publications

References 14 publications

Molecular Mechanisms of Pyrimidine Dimer Excision in Saccharomyces cerevisiae : Incision of Ultraviolet-Irradiated Deoxyribonucleic Acid In Vivo

Molecular Mechanisms of Pyrimidine Dimer Excision in Saccharomyces cerevisiae : Incision of Ultraviolet-Irradiated Deoxyribonucleic Acid In Vivo

Quantitative characterization of pyrimidine dimer excision from UV‐irradiated DNA (excision capacity) by cell‐free extracts of the yeast Saccharomyces cerevisiae

Repair of UV-irradiated plasmid DNA in a Saccharomyces cerevisiae rad3 mutant deficient in excision-repair of pyrimidine dimers

Contact Info

Product

Resources

About