Spontaneous mutation in lac I transgenic mice: a ... - Mutagenesis

Mutagenesis vol.13 no.2 pp.109-114, 1998

Spontaneous mutation in lac I transgenic mice: a comparison of tissues

Johan G. de Boer1-5, Scott Provost2, Nancy Gorelick3, Ken Tindall4 and Barry W. Glickman 'Centre for Environmental Health, University of Victoria, Victoria, BC, Canada V8W 2Y2, 2In Vitrogen Inc., Carlsbad, CA, USA, ^ e Procter and Gamble Company, Cincinnati, OH, USA and 4Narional Institute of Environmental Health Sciences, Research Triangle Park, NC, USA

The nature of spontaneous mutations in the lacl transgene of Big Blue® mice was determined in selected tissues. The mutant frequencies ranged from 2.5 x 10~5 to 7.1 X 10~5 for liver, spleen, bladder, stomach, kidney, bone marrow, lung and skin. We also determined the DNA sequence alterations in the mutants recovered from these tissues. In all tissues the predominant class of mutations was G:C—>A:T transitions, most of which occurred at 5'-CpG-3' dinucleotide sequences. Bladder, kidney and skin display the highest contribution of G:C—»A:T transitions. The second most common class of mutations was G:C—>T:A transverslons. All other base substitution classes contributed A:T A:T->G:C G:C-»T:A G:C->C:G A:T->T:A A:T->C:G + 1 Frameshift -1 Frameshift Deletions Insertions Complex Double substitution

Spleen

Exp. A 49 (59)°

Exp. B 52 (65)

Total 101 (124)

Exp. A 39(46)

Exp. B 51(57)

Total 90(103)

57.1 (78.6)b 2.0 18.4 (44.4) 2.0 6.1 2.0 2.0 8.2 2.0 0 0 0

40.4 (81.0) 7.7 11.5 (33.3) 5.8 3.8 3.8 3.8 15.4 7.7 0 0 0

48.5 (79.6) 5.0 14.9 (40.0) 4.0 5.0 3.0 3.0 11.9 5.0 0 0 0

38.5 (93.3) 12.8 23.1 (33.3) 5.1 2.6 5.1 2.6 0.0 5.1 2.6 0.0 2.6

42.1 (77.3) 5.9 15.7 (25.0) 7.8 5.9 3.9 0.0 5.9 9.8 2.0 0.0 0.0

41.1 (83.3) 8.9 18.9 6.7 4.4 4.4 1.1 3.3 7.8 2.2 0.0 1.1

Exp. A

Exp. B

Total

74.1 (83.7) 1.7 10.3 (33.3) 0.0 1.7 0.0 0.0 8.1 5.2 0.0 0.0 0.0

56.8 (95.2) 2.7 18.9 (57.1) 5.4 0.0 0.0 0.0 7.4 8.1 0.0 0.0 0.0

67.4 (87.5) 2.16 13.7 (46.2) 2.1 1.1 0.0

Bone marrow

G:C-»A:T A:T->G:C G:C->T:A G:C->C:G A:T->T:A A:T->C:G + 1 Frameshift -1 Frameshift Deletions Insertions Complex Double substitution

G:C->A:T A:T->G:C G:C-»T:A G:C->C:G A:T->T:A A:T->C:G + 1 Frameshift —1 Frameshift Deletions Insertions Complex Double substitution

Bladder

Exp. A 28 (33)

Exp. B 86(110)

28.6 (62.5) 7.1 28.6 (50.0) 3.6 7.1 14.3 0.0 3.6 3.6 0.0 0.0 3.6

45.3 (76.9) 10.5 11.6 (40.0) 4.7 4.7 0.0 5.8 8.1 4.7 2.3 2.3 0.0

Skin 37 (39)

Stomach 51 (59)

Kidney 60(81)

Liver 282 (348)

59.0 (87.0) 8.1 18.9 (37.5) 0.0 0.0 0.0 2.7 2.7 0.0 0.0 8.1 0.0

47.1 (83.3) 3.9 29.4 (40.0) 2.0 0.0 0.0 3.9 7.8 3.9 2.0 0.0 0.0

55.0(81.8) 1.7 20.0 (50.0) 6.7 3.3 1.7 1.7 3.3 3.3 3.3 0.0 0.0

48.9 (74.6) 6.0 18.4 (38.5) 3.9 2.8 3.2 1.8 7.1 3.2 1.8 0.7 2.1

Total 111 (143) 39 6 (76.9) 9.9 16.2 4.5 5.4 3.6 4.5 7.2 4.5 1.8 1.8 0.9

6.3 0.0 0.0 0.0

•The number of independent mutants. The number in parentheses is the number of mutants recovered. b The numbers in the mutation class rows are percentages. The numbers in parentheses are the percentage found at 5'-CpG-3' sites. •The numbers in the mutation class rows are percentages. The numbers in parentheses are the percentage found at 5'-CpG-3' sites.

found in liver, lung, brain and ovaries of female mice (Washington et ai, 1989). In addition, DNA repair activity may differ between cell types in an organ: O6-alkylDNA transferase activity varies significantly between the forestomach and corpus, with related tissues exhibiting intermediate values (Zaidi et al., 1993; Zaidi and O'Conner, 1995). On the basis of tissue-specific differences in DNA repair, proliferation or metabolism we may expect to observe variation in the types of mutation recovered from different tissues.

The main components of spontaneous mutation spectra in the lacl gene The pattern of spontaneous lacl mutation in transgenic mouse tissues is non-random. Transitions at G:C base pairs predominate and these occur most frequently at sites contained within 5'-CpG-3' dinucleotides (Table II). The cytosines within these sequences are often methylated. Methylated cytosines are prone to deamination, which results in conversion of 5-methylcytosine to thymine. Unlike uracil (the deamination product of cytosine), thymine is not 111

Downloaded from https://academic.oup.com/mutage/article-abstract/13/2/109/1021993/Spontaneous-mutation-in-lacI-transgenic-mice-a by guest on 28 September 2017

de Boer et al

removed by uracil-DNA glycosylase (Duncan and Miller, 1980). The resulting G:T mismatch will lead to a G:C->A:T transition, unless repaired by mammalian thymine-DNA glycosylase (Ehrlich et al, 1990) prior to DNA replication. The predominance of G:C—>A:T transitions at 5'-CpG-3' sites has also been reported by others for the lad gene recovered from transgenic Big Blue® mice (see for example Provost and Short, 1994; Sisk et al., 1994; Gorelick et al., 1995). This change does not predominate in the spectra of mutations recovered in the lad gene in Escherichia coli, where 5-methylcytosine occurs in 5'-CCAGG-3' sequences (Coulondre et al., 1978). The prevalence of mutations at 5'-CpG-3' sequences is also evident among mutations in various inherited human diseases. Examples include genes involved in hemophilia, Gaucher's disease (Choy et al., 1994) and factor IX (Siguret et al., 1988; Ketterling et al., 1994) and the p53 gene (Rideout et al., 1990). The second largest class of substitution mutations observed in all tissues (Table II) are G:C->T:A transversions. The source of these events is unknown, but they may reflect 8-oxoguanine (8-oxoG), a product of the reaction between reactive oxygen species and DNA. DNA synthesis past an 8-oxoG lesion may involve incorporation of an adenine opposite the 8-oxoG to produce a G:C—>T:A transversion. Moriya (1993) demonstrated that 8-oxoG in a single-stranded shuttle vector directs G—»T transversions (>78%) in simian kidney cells. Abasic sites may also generate G:C—>T:A transversions by insertion of adenine opposite the lesion (Sagher and Strauss, 1983). Distortions of the sugar-phosphate backbone are minimal for both 8-oxoG:G and 8-oxoG:A mispairs, but are sequence dependent (Poltev et al., 1993). G:C—»C:G transversion may be accounted for by replication of 8-oxoG:G mispairs. Accordingly, transversions at G:C base pairs would also be accounted for by oxidative damage. The fact that G:C—>C:G transversions are relatively rare may reflect the activities of OH8Gua glycosylase and OH8Gua endonuclease (Bessho et al., 1993), as well as an 8-oxo-dGTPase, which acts upon the dGTP nucleotide pool (Sakumi et al., 1993); all three activities have been identified in human cells. In E.coli MutY and MutM remove 8-oxoG lesions from DNA, greatly reducing transversion at G.C sites (Michaels et al., 1992). The remaining mutations found in the spontaneous spectra are mainly comprised of frameshifts, deletion and insertions. Approximately 50% of all frameshifts are found in runs of identical nucleotides, with deletions of single base pairs ~3-fold more frequent than additions. Deletions range from a few base pairs to ~80 bp. However, the Big Blue® system is capable of recovering longer deletions, as the longest deletion we have recovered in this system spans 800 bp of lad sequence (Stuart et al., unpublished results). Much larger deletions cannot be recovered because of bacteriophage packaging restrictions, the PCR amplification strategy and the fact that intergenic deletions cannot be recognized as such (Dycaico et al., 1994). Moreover, the lacl gene is exquisitely sensitive to base substitutions, which lowers the potential for deletion recovery. Mutational differences between tissues Organs develop from different cell layers during embryogenesis. The spleen, bone marrow and kidneys develop from the mesoderm layer, while the liver and the epithelial lining of the bladder, stomach, gut and lung develop from the endoderm. Skin and other epidermal cells are derived from the ectoderm

(Langman, 1973). Such differences may underly variations in metabolism and DNA repair and be reflected in the origin of spontaneous mutations. Table III shows the results of statistical analyses using the approach of Adams and Skopek (1987), performed to compare the distribution of mutation classes in the tissues studied here. The analysis also includes the contribution of G:C—»A:T transitions at CpG and non-CpG sites. With the exception of bladder, mutational spectra from the various tissues are very similar. In the case of bladder, however, the distribution of transitions at CpG sites was enhanced compared with that seen in other tissues. This may reflect differences in CpG methylation that occur in the bladder during development. Additionally, this difference may potentially reflect the lower pH (as low as 5) of urine (Kadlubar et al., 1991), which may result in a lowering of the intracellular pH in the cells lining the bladder. A reduced pH would enhance the rate of deamination (Wang et al, 1982) and, hence, account for the enhanced recovery of transitions found at 5'-CpG-3' dinucleotides in the bladder (Table II). While unique, the mutational spectrum in bladder is closest to that obtained from kidney (Table III), even though kidney and epithelial lining of the urinary bladder are not derived from the same embryonic tissue. We must point out, however, that the tissues examined in this study are not differentiated by tissue layer. The bladder and kidney, for example, are built from different tissue layers which may not be derived from the same embryonic stem cells. Bladder is also the only tissue that was isolated from female animals. This may also contribute to the difference in spectra between bladder and other tissues. The mutational spectrum in skin does not differ substantially from that seen in other tissues (Table UT). It is the only tissue in this survey that originates from the ectoderm. Different cell types have different enzymatic make-ups. For example, relatively high expression of enzymes involved in metabolic activation and deactivation of exogenous compounds, including the P450 cytochromes and various transferases (Rosenberg, 1991; Simpson et al, 1993; Kashfi et al, 1994), is found in the liver. Cells in the bone marrow also differ from other tissues due to the fact that they express high levels of myeloperoxidases. These differences in the levels of enzymes may be directly involved in the cell-specific response to exogenous compounds, but may have only minor consequences for spontaneous mutation, as indicated by the data presented here. The minor differences in spectra among these tissues may be partly due to differential methylation. The X. shuttle vector in Big Blue® mice is heavily methylated (Kohler et al, 1990) and this is probably also true of the lacl gene (Hogreve, unpublished data). Overall, the role of 5'-CpG-3' sites seems fairly consistent in all of the tissue types examined. Mutational spectra differences could reflect differences in the levels or the sites of methylation. The large contribution of G:C—>A:T transitions at 5'-CpG-3' sequences, however, may mask tissuespecific differences in other types of mutations. An examination of this issue would require the sequencing of a larger number of mutants or the construction of strains with a significantly reduced number of 5'-CpG-3' sequences. Such a lad gene has been constructed (Skopek et al, 1996). Strand specificity of mutations recovered at 5'-CpG-3' sequences The numbers of mutants recovered at the 5'-C and the 3'-G nucleotides (coding strand) in 5'-CpG-3' dinucleotide

112 Downloaded from https://academic.oup.com/mutage/article-abstract/13/2/109/1021993/Spontaneous-mutation-in-lacI-transgenic-mice-a by guest on 28 September 2017

Spontaneous mutation in lael transgenic mice: a comparison of tissues Table HI. P value analysis of comparison between tissues Liver Liver Lung Spleen Marrow Bladder Stomach Skin Kidney

0.62 0.51 0.68 0.016 0.62 0.40 0.81

Lung

Spleen

Marrow

Bladder

Stomach

Skin

Kidney

0.62

0.51 0.34

0.68 0.68 0.80

0.016 0.07 0.004 0.001

0.62 0.37 0.30 0.40 0.10

0.40 0.05 0.15 0.15 0.03 0.44

0.81 0.45 0.79 0.44 0.17 0.74 0.25

0.34 0.68 0.07 0.37 0.05 0.45

0.80 0.004 0.30 0.15 0.79

0.001 0.40 0.15 0.44

0.10 0.03 0.17

0.44 0.74

0.25

The calculation of P values was according to Adams and Skopek (1987), using the mutation classes shown in Table II with G:C-»A:T transitions separated into CpG and non-CpG groups.

Table IV. Strand specificity of C-»T transitions at 5'-CpG-3' dinucleotide sequences Tissue

C—»T (coding strand)

C->T (non-coding strand)

Liver Bone marrow Spleen Lung Stomach Kidney Skin Bladder

50 16 15 15 11 17 11 29

50 16 13 14 9 10 9 25

sequences of the coding strand of the lael gene are compared in Table IV (corrected for clonal expansion). C—>T transitions would reflect methylation of cytosine in the coding strand, while G-»A transitions would reflect methylation of cytosine in the non-coding strand. There is no evidence for strand specificity of potentially deamination-mediated transitions in these tissues, as the number of changes at the C and the G of 5'-CpG-3' sequences are very similar, with the possible exception of kidney, where a weak preference for the coding strand is noted. Overall, when applying a goodness-of-fit test on the two data sets (at C and at G) using the seven tissues as seven groups a P value of 0.84 is obtained (X2 = 2.7 for 6 degrees of freedom). This finding is consistent with the report that the integrated lael gene is heavily methylated and not expressed in mouse germ cells (Provost and Short, 1994). Comparison between experiments The statistical similarity between spectra of mutations recovered from different experiments for a given tissue highlights the reproducibility of these spectra when sufficient numbers of mutants are analyzed. The 'outlier' spectrum for bone marrow contains a relatively small number of mutants (28 independent mutants) and may indicate that the number of mutants required for analysis is larger. We note that the distribution and nature of mutations may vary between different experiments or laboratories. The reasons for these variations may rest in different sensitivities of color detection or the use of different animal strains, sexes or ages. However, analysis of 22 mutants with a faint blue phenotype (CM0 and CM1) recovered from control animals from various experiments indicates that there is no difference in spectrum (P > 0.9), using the Adams and Skopek (1987) statistical assay, when compared with mutations with all ranges of detectable colors (De Boer et al, submitted). Even frameshift and termination mutations may result in a light as well as a dark plaque color. This indicates that an inability to detect very faint blue mutant plaques, although it will underestimate

the mutant frequency, may not alter the mutant spectrum. Data collected before assay standardization is therefore not expected to change the spectra. In addition, there is no great difference in mutational spectra between the B6C3F1 and C57BL/6 lines, other than possibly a difference in the fraction of G:C—»A:T transitions found at 5'-CpG-3' dinucleotides in liver (De Boer et al, 1997). The utility of the transgenic system for mutation detection is greatly enhanced by the ability to examine mutational events at the DNA sequence level. This provides a unique opportunity to detect mutation at levels that only marginally increase the observed mutant frequency. For example, tris(l,2-dibromopropyl)phosphate increased the mutant frequency in the kidney of Big Blue® mice by only ~50%, however, an examination of the mutational spectrum revealed a significant increase in frameshift mutations (De Boer et al, 1996). In addition, DNA sequence analysis may provide insight into the mechanisms of mutagenesis. Spontaneous mutation has characteristic elements and this feature allows detection of changes in the spectrum of recovered mutations. The establishment of spontaneous spectra for various tissues is an important milestone for comparison of these induced spectra. Acknowledgements The authors would like to thank H.Erfte, J.Holcroft, D.Walsh and N.Hague for excellent technical support. This work was supported by a grant from the National Institutes of Environmental Health Sciences and the National Cancer Institute of Canada.

References Adams,W.T. and Skopek,T.R. (1987) Statistical test for the comparison of samples from mutational spectra. J. Mol. Bioi, 194, 391-396. Bessho.T, Tano.K., Kasai.H., Ohtsuka,E. and Nishimura,S. (1993) Evidence for two DNA repair enzymes for 8-hydroxyguanine (7,8-dihydro-8oxoguanine) in human cells. J. Biol Chem., 268, 19416-19421. Choy.F.Y., Wei.C, Applegarth.D.A. and McGillivray.B.C. (1994) DNA analysis of an uncommon missense mutation in a Gaucher disease patient of Jewish-Polish-Russian descent Am. J. Med Genet., 51, 156-160. Coulondre.C, MillerJ.H., Farabaugh.PJ. and Gilbert,W. (1978) Molecular basis of base substitution hotspots in Escherichia coli. Nature, YI4,775-780. De BoerJ.G. (1995) Software package for the management of sequencing projects using lael transgenic animals. Environ. Mol. Mutagen., 25,256-262. De BoerJ.G., MirsalisJ.C, Provost,G.S., Tindall,K.R. and Glickman.B.W. (1996) The spectrum of mutations in kidney, stomach and liver from lael transgenic mice, recovered after treatment with tris(2,3dibromopropyl)phosphate. Environ. Mol Mutagen., 28, 418-423. De BoerJ.G., Erfle.H., Walsh,D., HolcrofU-. ProvosUS., Rogers.B., Tindall,K.R. and Glickman.B.W. (1997) The Spectrum of spontaneous mutants in liver tissue of lael transgenic mice. Environ. Mol Mutagen., 30, 273-286. Duncan,B.K. and MillerJ.H. (1980) Mutagenic deamination of cytosine residues in DNA. Nature, 287, 560-561. Dycaico,MJ., Provost,G.S., KretzJ'.L., Ransom,S.L., MooresJ.C. and

113 Downloaded from https://academic.oup.com/mutage/article-abstract/13/2/109/1021993/Spontaneous-mutation-in-lacI-transgenic-mice-a by guest on 28 September 2017

de Boer et aL Short J.M. (1994) The use of shuttle vectors for mutation analysis in transgenic mice and rats. Mutat. Res., 307, 461—478. Ehrlich.M., Zhang.X.-Y. and Inamdar,N.M. (1990) Spontaneous deamination of cytosine and 5-methylcytosine residues in DNA and replacements of 5methylcytosine residues with cytosine residues. Mutat. Res., 238, 277-286. Gorelick.N.J., AndrewsJ., Gu,M. and Glickman.B.W. (1995) Mutational spectra in the lacl gene in skin from 7,12-dimethylbenz[a]anthracenetreated and untreated transgenic mice. Mol. Carcinogen., 24, 53-62. Gundersen.G., Kolsto.A.-B., Larsen.F. and Prydz.H. (1992) Tissue-specific methylation of a CpG island in transgenic mice Gene, 113, 207-214. Kadlubar.F.F, Dooley.K.L., Teitel.C.H., Roberts.D.W, Benson.R W., Butler.M.A., BaileyJ.R., YoungJ.F, Skipper,P.W. and Tannenbaum.S.R. (1991) Frequency of urination and its effects on metabolism, pharmacokinetics, blood hemoglobin adduct formauon, and liver and urinary bladder DNA adduct levels in beagle dogs given the carcinogen 4-aminobiphenyl. Cancer Res , 51, 4371-4377. Kashfi.K , Rimarachin.J A , Weksler.B.B. and Dannenberg.AJ (1994) Differential induction of glutathione S-transferase in rat aorta versus liver. Biochem. Pharmacol, 47, 1903-1907. Ketteriing.R.P, Vielhaber.E. and Sommer.S S. (1994) The rates of G C->TA and G:C—»C:G transversions at CpG dinucleotides in the human factor IX gene. Am. J. Hum. Genet., 54, 831-835. Kochanek.S., Toth.M , Dehmel.A , Renz.D. and Doerfler.W. (1990) Interindividual concordance of methylation profiles in human genes for tumor necrosis factors a and f3. Proc. Nad. Acad. Sci USA, 87, 8830-8834. Kohler.S.W., Provost,G.S., Kretz.P.L., Dycaico.M J., SorgeJ A. and ShorU.M (1990) Development of a short-term, in vivo mutagenesis assay: the effects of methylation on the recovery of a lambda phage shuttle vector from transgenic mice. Nucleic Acids Res , 18, 3007-3013. Kohler.S.W., Provost.G S , Fieck.A , Kretz.P.L., Bullock.W.O, SorgeJ.A., Putman.D.L. and ShortJ.M. (1991) Spectra of spontaneous and mutagenmduced mutations in the lacl gene in transgenic mice Proc. Nail. Acad Sci. USA, 88, 7958-7962. Kolachana.P., Subrahmanyam.V.V., Meyer.K.B., Zhang,L. and Smith.M.T. (1993) Benzene and its metabolites produce oxidative DNA damage in HL60 cells in vitro and in the bone marrow in vivo. Cancer Res., Si, 1023-1026 LangmanJ (1973) Medical Embryology, 2nd Edn. Williams & Wilkins Co., Baltimore, MD. Lee.Y.S., Lee.H.S., Park.M.K., Hwang.E.S., Park.E.M., Kasai.H. and Chung,M.H. (1993) Identification of 8-hydroxyguanine glycosylase activity in mammalian tissues using 8-hydroxyguanine specific monoclonal antibody Biochem. Biophys. Res. Commun., 196, 1545-1551. Lipkin,M. and Quastler.H (1962) Cell population kinetics in the colon of the mouse. J Clin. Invest., 41, 141-146. Michaels.M L., Cruz,C, Grollman.A.P. and MillerJ.H. (1992) Evidence that MutY and MutM combine to prevent mutations by an oxidatively damaged form of guanine in DNA Proc. Natl Acad. Sci USA, 89, 7022-7025. Mirsalis.J.C , Provost.G S., Matthews.C.D., Hamner.R.T, SchindlerJ E, O'Loughlin.K.G., MacGregorJ.T. and ShortJ.M (1993) Induction of hepatic mutations in lacl transgenic mice. Mutagenesis, 8, 265-271 Monya,M. (1993) Single-stranded shuttle phagemid for mutagenesis studies in mammalian cells' 8-oxoguamne in DNA induces targeted G:C—»TA transversions in simian kidney cells Proc. Natl. Acad. Sci. USA. 90, 1122-1126. Poltev.V.I., Smirnov.S.L., Issarafutdinova.O.V. and Lavery.R. (1993) Conformations of DNA duplexes containing 8-oxoguanine. J. Biomol. Struct. Dyn., 11, 293-301. Provost,G.S. and ShortJ.M. (1994) Characterization of mutations induced by ethylnitrosourea in seminiferous tubule germ cells of transgenic B6C3F1 mice. Proc. Natl. Acad. Sci. USA. 91. 6564-6568. RaffertyJ.A , Fan.C Y, Potter.P.M , Watson.A J.. Cawkwell.L . O'Conner.PJ. and Margison.G.P. (1992) Tissue-specific expression and induction of human O6-alkylguanine-DNA alkyltransferase in transgenic mice. Mol. Carcinogen., 6, 26-31. Rao.K S. and Loeb,L.A. (1992) DNA damage and repair in brain: relationship to aging. Mutat. Res., 275, 317-329. RideouuWM., Coetzee.G.A.. Olumi.A.F. and Jones.P.A. (1990) 5Methylcytosine as an endogenous mutagen in the human LDL receptor and p53 genes. Science, 249, 1288-1290. Rogers.B.J . Provost.G.S.. Young.R.R. Putman.D.L. and ShortJ.M. (1995) Intralaboratory optimization and standardization of mutant screening conditions used for a lambda/Zac/ transgenic mouse mutagenesis assay (I). Mutat. Res.. 327. 57-66 Rosenberg,D.W. (1991) Tissue-specific induction of the carcinogen inducible cytochrome P450 isoform, P4501A1, in colonic epithelium Arch Biochem. Biophys.. 284. 223-226. Sagher.D. and Strauss.B. (1983) Insertion of nucleotides opposite apurinic/

apyrimidinic sites in deoxyribonucleic acid during in vitro synthesis, uniqueness of adenine nucleotides. Biochemistry, 22, 4518—4526. Sakumi.K., Furuichi.M., Tsuzuki,T, Kakuma,T, Kawabata,S., Makj.H. and Sekiguchi,M. (1993) Cloning and expression of cDNA for a human enzyme that hydrolyzes 8-oxo-dGTP, a mutagenic substrate for DNA synthesis. J Biol Chem., 268, 23524-23530. Siguret,V., Amselem,S., Vidaud.M., Assouline,Z., Kerbiriou-Nabias.D., Pietu.G., Goossens.M., Larrieu.MJ., BahnalcB. and Meyer,D. (1988) Identification of a CpG mutation in the coagulation factor-IX gene by analysis of amplified DNA sequences. Br. J. Haematol., 70, 411^116. Simpson.E.R., Mahendroo.M.S Means.G.D., Kilgore.M.W, Corbin.CJ. and Mendelson.C.R. (1993) Tissue-specific promoters regulate aromatase cytochrome P450 expression / Steroid Biochem Mol. Biol., 44, 321-330. Sisk,S.C, Pluta.LJ., BondJ.A. and Recio.L. (1994) Molecular analysis of lacl mutants from bone marrow of B6C3F1 transgenic mice following inhalation exposure to 1,3-butadiene Carcinogenesis, 15, 471—477. Skopek,T.R., Mario.D.R., Kort,K L., MillerJ. and Pippert.T. (1996) Synthesis of a lacl gene analogue with reduced CpG content Mutat. Res., 349, 163-72. Wang,R.Y.-H, Kuo.K.C, Gehrke,C.W., Huang,L.-H. and Ehrlich.M (1982) Heat and alkali-induced deamination of 5-methylcytosine and cytosine residues in DNA. Biochim. Biophys. Ada, 697, 371-377. Washington.WJ., Dunn.W.CJ., Generoso,W.M. and Mitra,S. (1988) Tissuespecific variation in repair activity for 3-methyladenine in DNA in two stocks of mice. Mutat. Res., 207, 165-169. Washington.WJ., Foote,R S., Dunn.W.CJ., Generoso.W.M. and Mitra,S. (1989) Age-dependent modulation of tissue-specific repair activity for 3methyladenine and O6-methylguanine in DNA in inbred mice. Mech. Ageing Dev., 48, 43-52. Young.R.R., Rogers.BJ , Provost.G.S., ShortJ M. and Putman.D.L. (1995) Interlaboratory comparison: liver spontaneous mutant frequency from lambda//ac/ transgenic mice (Big Blue) (II). Mutat. Res., 327, 67-73. Zaidi.N H. and O'Conner.PJ (1995) Identification in rat stomach mucosae of a cell population characterized by a deficiency for the repan of O6methyldeoxyguanosine from DNA. Carcinogenesis, 16, 461—469. Zaidi.N.H., Potten.C S., Margison.G.P., Cooper.D.P. and O'Conner.PJ. (1993) Tissue and cell specific methylation, repair and synthesis of DNA in the upper gastrointestinal tract of Wistar rats treated with single doses of iV-methyl-Ap-nitro-A'-nitrosoguanidine. Carcinogenesis, 14, 1981-1990. Received on January 22, 1997; accepted on October I, 1997

114

Downloaded from https://academic.oup.com/mutage/article-abstract/13/2/109/1021993/Spontaneous-mutation-in-lacI-transgenic-mice-a by guest on 28 September 2017