Cell cycle regulation of DNA repair in normal and repair deficient human cells

The regulation of nucleotide excision repair and base excision repair by normal and repair deficient human cells was determined. Synchronous cultures of WI-38 normal diploid fibroblasts and Xeroderma pigmentosum fibroblasts (complementation group D) (XP-D) were used to investigate whether DNA repair pathways were modulated during the cell cycle. Two criteria were used: (1) unscheduled DNA synthesis (UDS) in the presence of hydroxyurea (HU) after exposure to UV light or after exposure to N-acetoxy-acetylaminofluorene (N-AcO-AAF) to quantitate nucleotide excision repair or UDS after exposure to methylmethane sulfonate (MMS) to measure base excision repair; (2) repair replication into parental DNA in the absence of HU after exposure to UV light. Nucleotide excision repair after UV irradiation was induced in WI-38 fibroblasts during the cell cycle reaching a maximum in cultures exposed 14–15 h after cell stimulation. Similar results were observed after exposure to N-AcO-AAF. DNA repair was increased 2–4-fold after UV exposure and was increased 3-fold after N-AcO-AAF exposure. In either instance nucleotide excision repair was sequentially stimulated prior to the enhancement of base excision repair which was stimulated prior to the induction of DNA replication. In contrast XP-D failed to induce nucleotide excision repair after UV irradiation at any interval in the cell cycle. However, base excision repair and DNA replication were stimulated comparable to that enhancement observed in WI-38 cells. The distinctive induction of nucleotide excision repair and base excision repair prior to the onset of DNA replication suggests that separate DNA repair complexes may be formed during the eucaryotic cell cycle.     

Characterization of a UV-resistant strain,UVr-10, established from a human clonal cell line,RSb, with high sensitivity to UV, 4NQO,MNNG and interferon

Characterization was performed of a UV-resistant variant strain, UV^r-10, derived from a human clonal cell line, RSb, with high sensitivity not only to the lethal effect of 254-nm far-ultraviolet (UV) irradiation but also to the effects of 4-nitroquinoline 1-oxide (4NQO) and N-methyl-N′-nitro-N-nitrosoguanidine (MNNG), and to the cell proliferation inhibition (CPI) effect of human leukocyte interferon (HuIFN-α) preparations.Colony-formation assays confirmed the increased resistance of UV^r-10 cells to both UV and 4NQO, but no increased resistance to MNNG. The marked recovery from the inhibition of the total cellular DNA synthesis of UV^r-10 cells, estimated by [methyl-³H]thymidine ([³H]dThd) uptake into the cellular DNA materials, was seen during 6 h after irradiation or 4NQO treatment even under the conditions without the recovery uptake into those of the parent RSb cells, but not during 6 h after MNNG treatment. Comparative studies on the activity of DNA repair synthesis between UV^r-10 and RSb cells, by measuring the extent of UV-, 4NQO- or MNNG-induced unscheduled DNA synthesis (UDS) and DNA repair replication, revealed an increased activity of UV^r-10 cells to UV and 4NQO but no significant increase of the activity to MNNG. These results suggest that increased DNA repair activities of a UV^r-10 cell line may account for its becoming resistant to the lethal effect of UV and 4NQO.Concerning the CPI effect of HuIFN-α, UV^r-10 cells showed increased resistance. Further, the DNA synthesis activity of UV^r-10 cells was not so inhibited by HuIFN-α exposure as that of RSb cells. However, HuIFN-α-exposed UV^r-10 cells showed more enhanced levels of activity of pppA(2′p5′A)_n synthetase (2–5A synthetase) than the exposed RSb, thus suggesting that HuIFN-α could exert enough intracellular effect even in UV^r-10 cells.The implication of the increased resistance of UV^r-10 cells to the effects of UV, 4NQO and HuIFN-α, but not to those of MNNG, is discussed.     

PHA刺激对淋巴细胞修复DNA损伤的影响

本文报道PHA刺激对淋巴细胞DNA修复的影响的实验结果。以254nm波长的UV照射细胞(30J/m~2)引起DNA损伤,以[~3H]-TdR掺入实验测定非程序DNA合成,用超微量法测定细胞的NAD~+含量,并以[~(35)S]-蛋氨酸掺入,聚丙烯酰胺凝胶电泳及放射自显影术测定蛋白质生物合成,其结果如下: (1)在被PHA转化的淋巴细胞内非程序DNA合成,随PHA刺激的时间加长而增高;PHA处理淋巴细胞42小时,合成的速率约增加4倍;(2)在转化的淋巴细胞内,非程序DNA合成及程序DNA合成都被N-乙基马来酰亚胺(一种DNA聚合酶α的抑制剂)抑制,表明在DNA修复过程中DNA聚合酶α可代替DNA聚合酶β发挥作用; (3)UV照射后,被PHA刺激的淋巴细胞内NAD~+含量大约减少43.2％,而对照淋巴细胞内NAD~+的含量只减少25％,似乎说明PHA刺激能促进淋巴细胞内的P-ADP-核糖化作用;(4)在受PHA刺激72小时的淋巴细胞内有多种蛋白质合成,这些细胞在UV照射后以含10μg/ml嘌呤霉素的培养基培养,则非程序DNA合成被明显抑制(P<0.01),这提示DNA修复是一需要蛋白质合成的过程。此外,在受UV照射后10-45小时的淋巴细胞内,诱导产生一种分子量大约34000道尔顿的蛋白质。 上述结果表明,当PHA使淋巴细胞从静止状态转化为增殖状态时,有多种酶被诱导。由于这些酶,如DNA聚合酶α及P-ADP-核糖聚合     

DNA damage and repair induced in vitro by nitrilotriacetic acid (NTA) in human lymphocytes

In cultured human lymphocytes we determined the ability of nitrilotriacetic acid (NTA) to inhibit DNA replication and to stimulate DNA repair synthesis (UDS), as well as to influence the UDS induced by UV irradiation. In phytohemagglutinin-stimulated lymphocytes a strong inhibition of DNA replication was induced by NTA concentrations above 10(-3) M, which was accompanied by a marked cell lethality, whereas at lower doses the incorporation of tritiated thymidine (3H-TdR) into DNA or treated cells was slightly increased in comparison to untreated cells. When, after NTA pretreatment, UDS was determined by scintillation spectrometry or autoradiography in unstimulated G0 lymphocytes, UV-irradiated or unirradiated, an increased incorporation of 3H-TdR was observed, positively correlated with the NTA doses. This effect was only partially due to the expansion of the intracellular TdR pool as a consequence of the stimulation of 3H-TdR uptake by NTA. Even after normalization of the scintillometric data by the radioactivities of the soluble nucleotide fraction, significant increase of DNA repair synthesis was detected after treatment with 7.5 x 10(-3)-10(-2) M NTA.     

A Comparison of Survival and Repair of Uv-Induced DNA Damage in Cultured Insect VERSUS Mammalian Cells

Survival and unscheduled DNA synthesis (UDS) were measured in a cultured insect cell line, TN-368, and a cultured mammalian cell line, V-79-4, following exposure to several fluences of ultraviolet light. TN-368 cells were approximately seven times more resistant to the lethal effects of UV than V-79 cells, as determined by colony formation. The amount of UDS per unit amount of DNA is about the same in both cell types 4 hr after 10–50 J/m² UV irradiations.     

X-ray dose-effect relationship on unscheduled DNA synthesis and spontaneous unscheduled DNA synthesis in mouse brain cells studied in vivo

Summary X-irradiation of the head of adult mice leads to DNA repair synthesis (unscheduled DNA synthesis, UDS) in non-proliferating cells of the brain as shown autoradiographically after injection of³H-thymidine and subsequent irradiation. The extent of UDS induced by one and the same X-ray dose varies between different cell types and also between different brain areas. Within the range of X-ray doses studied (2 to 100 Gy) a linear dose effect relationship was observed. No evidence of a saturation effect was found. The slopes of the regression lines for the dose effect relationship differ considerably for the different cell types. Two interesting correlations were found, if the present results were compared with other data in the literature: (i) There seems to be a correlation between the extent of UDS and radiosensitivity of the different cell types, the cells with low DNA repair synthetic rates being more radiosensitive. (ii) The extent of UDS of the different cell types correlates well with the extent of protein synthesis of the corresponding cell types. Apart from radiation induced UDS, spontaneous UDS was found to occur in sham-irradiated animals. The extent of spontaneous UDS also differs considerably between different cell types as well as between different brain areas. The increase of spontaneous UDS with increasing duration of immobilization of the animals during sham irradiation suggests a relationship between spontaneous UDS and stress.     

A new flow cytometric method to follow DNA gap filling during nucleotide excision repair of UVc-induced damage

BACKGROUND: Several methods have been developed for studying the kinetics of DNA repair after exposure of cells to ultraviolet (UV) light, such as conventional assays measuring unscheduled DNA synthesis (UDS). In this study, we have developed an accurate and rapid method to follow DNA gap filling during nucleotide excision repair (NER) in normal human fibroblasts (NHFs) in response to UV-induced damage. METHODS: After UVc irradiation, aphidicolin was added to the culture to hold repair patches open. This allowed an efficient incorporation of biotin-21-dUTP during an endogenous DNA repair synthesis that was detected by flow cytometry. RESULTS: We showed that the DNA gap filling after UVc irradiation in NHFs increased with time up to 10 h after irradiation and that no repair synthesis activity could be detected in XP-A fibroblasts. Furthermore, this activity was UVc dose dependent up to 20 J/m2. These results correlated well with those of the UDS assay. Interestingly, addition of aphidicolin at different time points after UVc irradiation, thus allowing endogenous repair synthesis in the absence of biotin-21-dUTP, demonstrated that the response of the NER system occurred extremely rapidly after irradiation. CONCLUSIONS: This method may be a reliable and simple alternative to other techniques measuring UDS. Practical advantages include the rapidity of the method, no need for radioactivity, and the possibility to use a second and/even a third flow marker to analyse cell cycle and heterogeneous cell populations concomitantly.     

Cytological evidence for DNA chain elongation after UV irradiation in the S phase

Human cells irradiated with UV light synthesize lower molecular weight DNA than unirradiated cells. This reduction in molecular weight is greater in xeroderma pigmentosum (XP) cells than in normal cells. The molecular weight of DNA is further reduced by the addition of caffeine to XP cells. By several hours after irradiation, DNA fragments are barely detectable. Cells from excision-proficient and excision-deficient XP patients were studied autoradiographically to produce cytological evidence of DNA chain elongation. Replicate cultures with and without caffeine were synchronized and irradiated with UV light during the S phase. Caffeine was removed in G₂, and the cells were labeled with ³H-thymidine. Results showed significantly increased labeling during G₂ of excision-deficient XP cells. Labeling was dependent on both time of irradiation and presence of caffeine. The XP variant cells had no increase in labeling for any irradiation time.Published with the approval of the Director of the West Virginia Agricultural Experiment Station as Scientific Paper No. 1608. Supported by N.I.C. Grant TO1CA05170-10.     

Efficacy of UV Irradiation in Inactivating Cryptosporidiumparvum Oocysts

To evaluate the effectiveness of UV irradiation in inactivating Cryptosporidium parvum oocysts, the animal infectivities and excystation abilities of oocysts that had been exposed to various UV doses were determined. Infectivity decreased exponentially as the UV dose increased, and the required dose for a 2-log₁₀ reduction in infectivity (99% inactivation) was approximately 1.0 mWs/cm² at 20°C. However, C. parvum oocysts exhibited high resistance to UV irradiation, requiring an extremely high dose of 230 mWs/cm² for a 2-log₁₀ reduction in excystation, which was used to assess viability. Moreover, the excystation ability exhibited only slight decreases at UV doses below 100 mWs/cm². Thus, UV treatment resulted in oocysts that were able to excyst but not infect. The effects of temperature and UV intensity on the UV dose requirement were also studied. The results showed that for every 10°C reduction in water temperature, the increase in the UV irradiation dose required for a 2-log₁₀ reduction in infectivity was only 7%, and for every 10-fold increase in intensity, the dose increase was only 8%. In addition, the potential of oocysts to recover infectivity and to repair UV-induced injury (pyrimidine dimers) in DNA by photoreactivation and dark repair was investigated. There was no recovery in infectivity following treatment by fluorescent-light irradiation or storage in darkness. In contrast, UV-induced pyrimidine dimers in the DNA were apparently repaired by both photoreactivation and dark repair, as determined by endonuclease-sensitive site assay. However, the recovery rate was different in each process. Given these results, the effects of UV irradiation on C. parvum oocysts as determined by animal infectivity can conclusively be considered irreversible.     

The Dampening of DNA Replication Cycles after X-Irradiation or Ultraviolet Light Irradiation

Escherichia coli strains 15T^- (555-7) and B/r were grown in the presence of thymine-¹⁴C to label all DNA. The ability of these parental DNA's to undergo cycles of replication subsequent to cellular irradiation with either X-ray or ultraviolet light (UV) was followed with density labels. Exposed cells were shifted into the density medium at times which were approximately multiples of normal rounds of DNA replication. A portion of the parental DNA, replicated semiconservatively once during an initial cycle following UV or X-irradiation in E. coli, failed to replicate again within the time studied. The time course of semiconservative parental DNA replication is altered.     

Low-Pressure UV Inactivation and DNA Repair Potential of Cryptosporidium parvum Oocysts

Because Cryptosporidium parvum oocysts are very resistant to conventional water treatment processes, including chemical disinfection, we determined the kinetics and extent of their inactivation by monochromatic, low-pressure (LP), mercury vapor lamp UV radiation and their subsequent potential for DNA repair of UV damage. A UV collimated-beam apparatus was used to expose suspensions of purified C. parvum oocysts in phosphate-buffered saline, pH 7.3, at 25°C to various doses of monochromatic LP UV. C. parvum infectivity reductions were rapid, approximately first order, and at a dose of 3 mJ/cm² (=30 J/m²), the reduction reached the cell culture assay detection limit of ~3 log₁₀. At UV doses of 1.2 and 3 mJ/cm², the log₁₀ reductions of C. parvum oocyst infectivity were not significantly different for control oocysts and those exposed to dark or light repair conditions for UV-induced DNA damage. These results indicate that C. parvum oocysts are very sensitive to inactivation by low doses of monochromatic LP UV radiation and that there is no phenotypic evidence of either light or dark repair of UV-induced DNA damage.     

Comparative induction of unscheduled DNA synthesis by physical and chemical agents in non-proliferating primary cultures of rat hepatocytes

The incorporation of [³H]thymidine into DNA due to unscheduled DNA synthesis (UDS) induced by N-OH-2-acetylaminofluorene (N-OH-AAF), aflatoxin B₁ (AFB₁), ethyl methanesulfonate (EMS) and ultra-violet light was quantitated by autoradiography and by scintillation spectrometry on acid precipitable macromolecules or DNA insolated by isopycnic banding in cesium chloride (CsCl). Dose-dependent increases in UDS due to N-OH-AAF and AFB₁ treatment were found. Only 2-fold increases at the highest dose levels were found, however, when incorporated [³H]thymidine was quantitated by scintillation spectrometry. Seven, 11, and 25-fold increases in UDS induced by AFB₁, N-OH-AAF and ultra-violet light, respectively, were found when incorporated [³H]thymidine was quantitated by autoradiography, indicating a high sensitivity for detecting ‘long patch’ repair by this technique. Scintillation spectrometry was completely ineffective in detecting EMS-induced UDS, whereas autoradiography demonstrated a small, but significant induction in [³H]thymidine incorporation at high dose levels. The non-proliferative nature of the primary hepatocyte prohibits the uniform radioactive prelabeling of DNA, necessary in other techniques, for the detection of ‘short patch’ repair induced by compounds such as EMS. Therefore, the sensitivity of the primary cultured rat hepatocyte in conjunction with UDS for detecting DNA damage caused by mutagens and carcinogens which induce ‘short patch’ repair may be limited to the autoradiographic analysis of the unscheduled incorporation of [³H]thymidine.     

Phenotypic heterogeneity within the first complementation group of UV-sensitive mutants of Chinese hamster cell lines

A DNA-repair mutant was characterized that has the extraordinary and interesting properties of extreme sensitivity to UV killing combined with a high level of nucleotide excision repair. The mutant V-H1 isolated from the V79 Chinese hamster cell line appeared very stable, with a reversion frequency of about 3.5 × 10⁻⁷. Genetic complementation analysis indicates that V-H1 belongs to the first complementation group of UV-sensitive Chinese hamster ovary (CHO) mutants described by Thompson et al. (1981). This correponds with data on cross-sensitivity and mutation induction after UV irradiation published by this group. Surprisingly, the mutant V-H1 shows only slightly reduced (to ∼ 70%) unscheduled DNA synthesis (UDS) after UV exposure, while the other two mutants of this complementation group are deficient in UDS after UV. In agreement with the high residual UDS, in V-H1 also the amount of repair replication in response to UV treatment is relatively high (∼ 50%). It has also been shown that the incision step of the nucleotide excision pathway takes place in V-H1 (with a lower rate than observed in wild-type cells), whereas another mutant (UV5) of the same complementation group is deficient in incision.This heterogeneity within the first complementation group indicates that the repair gene of this complementation group may have more than one functionally domain or that the gene is not involved in the incision per se but is involved in e.g. preferential repair of active genes.     

DNA Repair in Potorous tridactylus

The DNA synthesized shortly after ultraviolet (UV) irradiation of Potorous tridactylis (PtK) cells sediments more slowly in alkali than that made by nonirradiated cells. The size of the single-strand segments is approximately equal to the average distance between 1 or 2 cyclobutyl pyrimidine dimers in the parental DNA. These data support the notion that dimers are the photoproducts which interrupt normal DNA replication. Upon incubation of irradiated cells the small segments are enlarged to form high molecular weight DNA as in nonirradiated cells. DNA synthesized at long times (~ 24 h) after irradiation is made in segments approximately equal to those synthesized by nonirradiated cells, although only 10-15% of the dimers have been removed by excision repair. These data imply that dimers are not the lesions which initially interrupt normal DNA replication in irradiated cells. In an attempt to resolve these conflicting interpretations, PtK cells were exposed to photoreactivating light after irradiation and before pulse-labeling, since photoreactivation repair is specific for only one type of UV lesion. After 1 h of exposure ~ 35% of the pyrimidine dimers have been monomerized, and the reduction in the percentage of dimers correlates with an increased size for the DNA synthesized by irradiated cells. Therefore, we conclude that the dimers are the lesions which initially interrupt DNA replication in irradiated PtK cells. The monomerization of pyrimidine dimers correlates with a disappearance of repair endonuclease-sensitive sites, as measured in vivo immediately after 1 h of photoreactivation, indicating that some of the sites sensitive to the repair endonuclease (from Micrococcus luteus) are pyrimidine dimers. However, at 24 h after irradiation and 1 h of photoreactivation there are no endonuclease-sensitive sites, even though ~ 50% of the pyrimidine dimers remain in the DNA. These data indicate that not all pyrimidine dimers are accessible to the repair endonuclease. The observation that at long times after irradiation DNA is made in segments equal to those synthesized by nonirradiated cells although only a small percentage of the dimers have been removed suggests that an additional repair system alters dimers so that they no longer interrupt DNA replication.     

DNA repair investigations in nine Italian patients affected by trichothiodystrophy.

Trichothiodystrophy (TTD) is a rare autosomal recessive disorder characterized by brittle hair, mental and growth retardation, peculiar face, ichthyosis, and in 20% of the reported cases photosensitivity. Cellular photosensitivity due to the same genetic defect present in xeroderma pigmentosum group D (XP-D) has been described in several patients. Nine patients with clinical symptoms diagnostic for TTD have been identified in Italy to date. We report the results of DNA repair investigations performed in cultured fibroblasts from these patients and 8 TTD parents. Survival, DNA repair synthesis and RNA synthesis following UV irradiation were all normal in the 8 TTD heterozygous cell strains. Among the 9 TTD-affected individuals, normal cellular UV sensitivity was observed in the 2 patients without signs of clinical photosensitivity. In contrast, the other 7 TTD cell strains showed a notable reduction in UV-induced DNA repair synthesis (UDS) levels, ranging between 40% and 5-15% of normal values. Complementation analysis indicated that in the repair-deficient TTD cell strains the genetic defect is the same as that present in XP-D cells. The biochemical heterogeneity of the XP-D defect in TTD patients characterized by different degrees of defective UDS results in different patterns of response to the killing effect of UV light in non-proliferating cells.     

Water- and temperature-dependence of DNA damage and repair in the fruticose lichen Cladonia arbuscula ssp. mitis exposed to UV-B radiation

The induction of cyclobutane pyrimidine dimers (CPDs) by ultraviolet‐B radiation (UV‐B, 280–315 nm) and repair mechanisms were studied in the lichen Cladonia arbuscula ssp. mitis exposed to different temperatures and water status conditions. In addition, the development and repair of CPDs were studied in relation to the different developmental stages of the lichen thallus podetial branches. Air‐dried lichen thalli exposed to UV‐B radiation combined with relatively high visible light (HL, 800 μmol m^?2 s^?1; 400–700 nm) for 7 days showed a progressive increase of CPDs with no substantial repair, although HL was present during and after irradiation with UV‐B. Fully hydrated lichen thalli, that had not been previously exposed to UV‐B radiation for 7 days, were given short‐term UV‐B radiation treatment at 25°C, and accumulated DNA lesions in the form of CPDs, with repair occurring when they were exposed to photoreactivating conditions (2 h of 300 μmol m^?2 s^?1, 400–700 nm). A different pattern was observed when fully hydrated thalli were exposed to short‐term UV‐B radiation at 2°C, in comparison with exposure at 25°C. High levels of CPDs were induced at 2°C under UV‐B irradiation, without significant repair under subsequent photoreactivating light. Likewise, when PAR (300 μmol m^?2 s^?1) and UV‐B radiation were given simultaneously, the CPD levels were not lowered. Throughout all experiments the youngest, less differentiated parts of the lichen thallus – namely ‘tips’, according to our arbitrary subdivision – were the parts showing the highest levels of CPD accumulation and the lowest levels of repair in comparison with the older thallus tissue (‘stems’). Thus the experiments showed that Cladonia arbuscula ssp. mitis is sensitive to UV‐B irradiation in the air‐dried state and is not able to completely repair the damage caused by the radiation. Furthermore, temperature plays a role in the DNA damage repairing capacity of this lichen, since even when fully hydrated, C. arbuscula ssp. mitis did not repair DNA damage at the low temperatures.     

Reduced DNA repair in progeria cells and effects of gamma-ray irradiation on UV-induced unscheduled DNA synthesis in normal and progeria cells

A reduction in the amount of UV-induced unscheduled DNA synthesis (UDS), and reduced cell survival and host-cell reactivation against UV exposure in Hutchinson-Gilford progeria syndrome cell strains were shown. UV-induced UDS in 4 progeria cell strains was 33-50% of the normal level. A similar reduction in the UV-induced UDS in normal cells was caused by gamma-ray irradiation to the cells before UV irradiation. The dose of gamma-rays required to cause a reduction in UDS of normal cells to the level of progeria cells was 40 Gy and the reduction was reversible after 2 days. In progeria cells, gamma-ray irradiation further reduced UDS with a lower gamma-ray dose required than in normal cells, and the reduction was also reversible but with less relative recovery than in normal cells. The presence of a 'built-in' defect in progeria cells responsible for the reduced DNA-repair capacity was suggested, and such defect may share a common mechanism with the reduction of UV-induced UDS in normal cells caused by gamma-ray irradiation.     

Cellular and genetic studies in three UV-sensitive Chinese hamster mutants

Three UV sensitive (UV^s) mutants (CHO43RO, CHO423PV, CHO30PV), characterized by different levels of reduction in their ability to perform unscheduled DNA synthesis (UDS), were analysed for spontaneous and UV-induced frequency of chromosomal aberrations and for sensitivity to alkylating agents. The baseline frequency of chromosomal aberrations was in the normal range, whereas after UV irradiation a positive correlation between the degree of UV sensitivity and the rate of chromosomal breakage was observed. Survival experiments after mutagen exposure indicated that the UV^s clones are characterized by different levels of hypersensitivity to bifunctional alkylating agents whereas the sensitivity to monofunctional alkylating agents is in the normal range. Genetic analysis performed by measuring the survival after UV in hybrids produced by fusing UV^s cells with wild-type or UV^s cells belonging to the six Chinese hamster complementation groups, indicated that the three clones carry recessive mutations and belong to c.g. 2. These findings suggest that defects in the same gene may result in different degrees of phenotypic alterations.Abbreviations CG complementation group - EMS ethyl methane sulfonate - MMS methyl methane sulfonate - MMC mitomycin C - UV ultraviolet - UDS unscheduled DNA synthesis     

Excision repair of mouse and human fibroblast cells, and a factor affecting the amount of UV-induced unscheduled DNA synthesis

Excision-repair ability and the amount of unscheduled DNA synthesis (UDS) after UV irradiation of fibroblast cells (in vitro passage 5) from C57BL mouse embryos were compared with those of human skin fibroblast cells. UDS in the mouse cells was approximately 75% of that in the human cells, although the disappearance of T4 endonuclease-V-susceptible sites and the accumulation of single-strand breaks in the mouse cell DNA indicated that the excision-repair capacity of the mouse cells was 20-35% of that in the human cells. This apparent discrepancy was ascribed to the difference in intracellular dTTP pool size, which was approximately twice as large in the human cells as in the mouse cells. UDS may not be suitable as a quantitative measure of excision repair when comparing the cells from different species.