CHO mutant UV61 removes (6–4) photoproducts but not cyclobutane dimers

Abstract

The CHO mutant UV61 was previously assigned to complementation group 6 of UV-sensitive rodent cell mutants. UV61 is less sensitive to killing by UV radiation than mutants such as UV5, which is highly defective in the incision process that acts on UV-induced lesions. The D37 for cell survival is approximately 4 J/m2 for UV61, compared with 10 J/m2 for the parental AA8 line and approximately 2 J/m2 for UV5. Similarly, mutation induction at the hprt and aprt loci shows an intermediate response to UV61. In a post-replication recovery assay, the kinetics of maturation of pulse-labelled nascent DNA were normal after UV irradiation in UV61. Data from alkaline elution and alkaline unwinding assays showed that the rates of break accumulation and resealing, measured 0-120 min after irradiation, were also normal in the mutant. This repair incision correlated with the rapid, normal removal of pyrimidine(6-4)pyrimidone photoproducts in UV61 measured using a radioimmunoassay that is specific for this class of damage. In contrast, after exposure to 10 or 15 J/m2, no detectable removal of cyclobutane dimers from DNA was found in UV61 while AA8 cells removed 32% by 24 h. We suggest that the mutation in UV61 specifically lowers the affinity of a repair protein for cyclobutane dimers, which are also inefficiently removed from the bulk DNA of normal CHO cells. The resistance of UV61 to killing by the direct acting chemical 7-bromomethylbenz[a]anthracene was only slightly greater than that of UV5, indicating defective repair of bulky chemical adducts in addition to cyclobutane dimers.(ABSTRACT TRUNCATED AT 250 WORDS)

ERCC6, a member of a subfamily of putative helicases, is involved in Cockayne's syndrome and preferential repair of active genes

“…Presumably, as with allele a, (Troelstra et al, 1990). b Thompson et al (1988a). c CSlAN.SS.G2 (unpublished data).…”

Repair by human cell extracts of single (6-4) and cyclobutane thymine-thymine photoproducts in DNA.

“…One possibility is that the nucleotide excision repair system may recognize or incise each with significantly different efficiencies. Alternatively, there may be two partially redundant repair systems, only one ofwhich efficiently removes photoproducts (14,15). lesions might also be preferentially repaired if they are more accessible in chromatin to the repair complex, a possibility supported by evidence that (6-4) photoproducts (unlike cyclobutane dimers) form with higher yield in linker regions of nucleosomes than in the core (16,17).…”

Increased UV resistance of a xeroderma pigmentosum revertant cell line is correlated with selective repair of the transcribed strand of an expressed gene.

“…Cells in the classical XP complementation groups and the majority of cellular mutants that are defective in repair of UV-induced DNA damage are defective in repair of both lesions. However, a few rodent and human cell mutants that have altered repair capacities for only one of these photoproducts have been identified (6,13,20,27). These mutants should be useful for evaluating the respective biological roles of the photoproducts and may also provide insight into the mechanism of excision repair in mammalian cells.…”

“…These studies resulted in conflicting conclusions. For example, the Chinese hamster ovary (CHO) cell mutant UV61 efficiently removes 6-4 PPs but not CPDs when repair is assayed as an average over the entire genome (27). The UV resistance of UV61 cells is intermediate between that of the parental CHO cells and that of mutants that are deficient in repair of both photoproducts (21,27).…”