Summary
The centromere is the region within a chromosome that is required for proper segregation during mitosis and meiosis. Lesions in this sequence represent a unique type of damage, as loss of function could result in catastrophic loss of the genetic material of an entire chromosome. We have measured the induction by ultraviolet (UV) light of pyrimidine dimers in a 2550-bp restriction fragment that includes the centromere region of chromosome III in Saccharomyces cerevisiae. Yeast cells were exposed to ultraviolet light, cellular DNA was gently extracted, and subsequently treated with a UV-specific endonuclease to cleave all pyrimidine dimers. The sites of UV-specific nuclease scission within the centromere were determined by separating the DNA according to molecular weight, transferring the fragments to nitrocellulose, and hybridizing to a radiolabeled 624-bp fragment homologous to the centromere DNA from chromosome III. Several hotspots were identified in chromatin DNA from cells, as well as in irradiated deproteinized DNA. Double strand damage due to closely opposed pyrimidine dimers was also observed. At biological doses (35% survival) there are approximately 0.1 to 0.2 pyrimidine dimers per centromere. These dimers are efficiently repaired in the centromere and surrounding region.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Bloom KS, Carbon J (1982) Highly ordered structure in chromosomes and small circular minichromosomes. Cell 29:305–316
Bloom KS, Fitzgerald-Hayes M, Carbon J (1983) Structural analysis and sequence organization of yeast centromeres. Cold Spring Harbor Symp Quant Biol 47:1175–1185
Bloom KS, Amaya E, Carbon J, Clarke L, Hill A, Yeh E (1984) Chromatin conformation of yeast centromeres. J Cell Biol 99:1559–1568
Bohr VA, Smith CA, Okumoto DS, Hanawalt PC (1985) DNA repair in an active gene: Removal of pyrimidine dimers from the DHRF gene of CHO cells is much more efficient than that in the genome overall. Cell 40:359–369
Carrier W, Setlow RB (1970) Endonuclease from Micrococcus luteus which has activity toward ultraviolet-irradiated deoxyribonucleic acid; purification and properties. J Bacteriol 102:178–186
Clarke L, Carbon J (1980) Isolation of a yeast centromere and construction of functional small circular chromosomes. Nature 287:504–509
Clarke L, Carbon J (1983) Genomic substitutions of centromeres in Saccharomyces cerevisiae. Nature 305:23–28
DiCaprio L, Cox SB (1981) The effect UV-irradiation on the molecular weight of pre-existing and newly-synthesized DNA. Mutat Res 82:69–85
Esposito MS, Maleas DT, Bjornstad KA, Bruschi CV (1982) Simultaneous detection of changes in chromosome number, gene conversion and intergenic recombination during mitosis of Saccharomyces cerevisiae: Spontaneous and ultraviolet light induced events. Curr Genet 6:5–11
Evans DH, Linn S (1984) Excision repair of pyrimidine dimers from simian virus 40 minichromosomes in vitro. J Biol Chem 259:10252–10259
Fitzgerald-Hayes M, Clarke L, Carbon J (1982) Nucleotide sequence comparisons and functional analysis of yeast centromere DNAs. Cell 29:235–244
Forte MA, Fangman WL (1976) Naturally occurring cross-links in yeast chromosomal DNA. Cell 8:425–431
Gaudet A, Fitzgerald-Hayes M (1987) Alterations in the A+T-rich region of CEN3 affect centromere function in Saccharomyces cerevisiae. Mol Cell Biol 7:68–75
Hieter P, Pridmore D, Hegemann JH, Thomas M, Davis RW, Philippsen P (1985) Functional selection and analysis of yeast centromeric DNA. Cell 42:913–921
Lam LH, Reynolds R (1986a) Bifilar enzyme-sensitive sites in ultraviolet-irradiated DNA are indicative of closely opposed cyclobutyl pyrimidine dimers. Biophys J 50:307–317
Lam LH, Reynolds R (1986b) A sensitive, enzymatic assay for the detection of closely opposed pyrimidine dimers induced in human diploid fibroblasts. Mutat Res 166:187–198
Leblon G, Zickler D, Lebilcot S (1986) Most UV-induced reciprocal translocations in Sordaria macrospora occur in or near the centromere regions. Genetics 112:183–204
McGrew J, Diehl B, Fitzgerald-Hayes M (1986) Single base-pair mutations in centromere element III cause aberrant chromosome segregation in Saccharomyces cerevisiae. Mol Cell Biol 6:530–538
Mortimer RK, Schild D (1985) Genetic map of Saccharomyces cerevisiae, Edition 9. Microbiol Rev 49:181–212
Ng R, Cumberledge S, Carbon J (1986) Structure and function of centromeres. In: Hicks J (ed) Yeast Cell Biology. Alan R. Liss, New York, pp 225–239
Panzeri L, Landoia L, Stotz A, Philippsen P (1985) Role of conserved sequence elements in yeast centromere DNA. EMBO J 4:1867–1874
Parry JM, Sharp D, Tippins RS, Parry EM (1979) Radiation-induced mitotic and meiotic aneuploidy in the yeast Saccharomyces cerevisiae. Mutat Res 61:37–55
Prakash L (1981) Characterization of postreplication repair in Saccharomyces cerevisiae and effects of rad6, rad18 and rad52 mutations. Mol Gen Genet 184:471–478
Resnick MA (1970) Sunlight-induced killing in Saccharomyces cerevisiae. Nature 226:377–378
Resnick MA, Bloom K (1987) Lessons learned from yeast: A molecular and genetic analysis of centromere function. In: Vig BV, Sandberg A (eds) Aneuploidy: Incidence and Etiology. Alan R Liss, New York, pp 395–413
Resnick MA, Setlow JK (1972) Repair of pyrimidine dimer damage induced in yeast by ultraviolet light. J Bacteriol 109:979–986
Resnick MA, Boyce J, Cox B (1981) Postreplication repair in Saccharomyces cerevisiae. J Bacteriol 146:285–290
Resnick MA, Stasiewicz S, Game JC (1983) Meiotic DNA metabolism in wild-type and excision-deficient yeast following UV exposure. Genetics 104:583–601
Resnick MA, Sugino A, Nitiss J, Chow T (1984) DNA polymerases, deoxyribonucleases, and recombination during meiosis in Saccharomyces cerevisiae. Mol Cell Biol 4:2811–2817
Sedgwick SG (1976) Misrepair of overlapping daughter strand gaps as a possible mechanism for UV induced mutagenesis UVR strains of Escherichia coli: A general model for induced muta-genesis by misrepair (SOS repair) of closely spaced DNA lesions. Mutat Res 41:185–200
Unrau R, Wheatcroft R, Cox BS (1972) Postreplication repair in Saccharomyces cerevisiae. Biochem Biophys Acta 269:311–321
Wheatcroft R, Cox B, Haynes RH (1975) Repair of UV-induced DNA damage and survival in yeast. I. Dimer excision. Mutat Res 30:209–218
Williams JI, Friedberg EC (1979) Deoxyribonucleic acid excision repair in chromatin after ultraviolet irradiation of human fibroblasts in culture. Biochemistry 18:3965–3972
Yeh E, Carbon J, Bloom K (1986) Tightly centromere-linked gene (SPO15) essential for meiosis in the yeast Saccharomyces cerevisiae. Mol Cell Biol 6:158–167
Author information
Authors and Affiliations
Additional information
Communicated by B.J. Kilbey
Rights and permissions
About this article
Cite this article
Resnick, M.A., Westmoreland, J., Amaya, E. et al. UV-induced damage and repair in centromere DNA of yeast. Mol Gen Genet 210, 16–22 (1987). https://doi.org/10.1007/BF00337753
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF00337753