Restriction enzyme-induced DNA double-strand breaks as a model system for cellular responses to DNA damage.
To learn more about cellular responses to DNA double-strand breakage, we used three methods to assay cellular damage after treatment with a restriction enzyme that causes DNA double-strand breaks by cleaving at specific recognition sites in the DNA. Chinese hamster ovary cells were treated with increasing doses of Pvu II and studied for double-strand breakage, chromosomal aberration yield, and cell survival. The yield of DNA double-strand breaks, as measured by pulsed-field gel electrophoresis, increased at concentrations up to 500 units and saturated thereafter. The maximum yield of metaphase cells showing aberrant chromosomes was reached at 100 units and stayed constant up to 1,000 units. Although exchange-type aberrations saturated at approximately 4.5 per cell at 100 units, deletion-type aberrations appeared to increase at concentrations up to 500 units. Cell survival, as measured by colony-forming ability after Pvu II treatment, saturated at 100 units. The observed dose-response data are probably due to the saturation of accessible Pvu II cleavage sites within the cell. These data indicate that restriction enzymes induce the same DNA-damaging effects as many of the agents used in cancer treatment. Because the primary DNA lesion induced by restriction enzymes is known, they provide a unique opportunity to understand cellular responses to DNA damage and repair.
Yates
B L
BL
Laboratory of Radiobiology and Environmental Health, University of California, San Francisco 94143-0750.
Valcarcel
E R
ER
Morgan
W F
WF
D000818:Animals; D016466:CHO Cells; D002470:Cell Survival / Q000187:drug effects; D002869:Chromosome Aberrations; D006224:Cricetinae; D004247:DNA / Q000187:drug effects*; D004249:DNA Damage; D015252:Deoxyribonucleases, Type II Site-Specific / Q000494:pharmacology*; D008957:Models, Genetic
D004247:DNA; C025630:CAGCTG-specific type II deoxyribonucleases; D015252:Deoxyribonucleases, Type II Site-Specific