A Mechanism by which Ergosterol Inhibits the Promotion of Bladder Carcinogenesis in Rats.

We previously showed that ergosterol has an inhibitory effect on bladder carcinogenesis. In this study, we aimed to elucidate the molecular mechanism by which ergosterol inhibits bladder carcinogenesis using a rat model of N-butyl-N-(4-hydroxybutyl)nitrosamine-induced bladder cancer. The messenger ribonucleic acid (mRNA) expression level of the cell cycle-related gene cyclin D1 and inflammation-related gene cyclooxygenase-2 in bladder epithelial cells was significantly increased in the carcinogenesis group compared with the control group. In contrast, in ergosterol-treated rats, these increases were significantly suppressed. Ergosterol did not affect the plasma testosterone concentration or the binding of dihydrotestosterone to androgen receptor (AR). The mRNA expression levels of 5α-reductase type 2 and AR were higher in the carcinogenesis group than in the control group but were significantly decreased by ergosterol administration. These results suggest that ergosterol inhibits bladder carcinogenesis by modulating various aspects of the cell cycle, inflammation-related signaling, and androgen signaling. Future clinical application of the preventive effect of ergosterol on bladder carcinogenesis is expected.

Life-Shortening Effect of Chronic Low-Dose-Rate Irradiation in Calorie-Restricted Mice.

Calorie restriction is known to influence several physiological processes and to alleviate the late effects of radiation exposure such as neoplasm induction and life shortening. However, earlier related studies were limited to acute radiation exposure. Therefore, in this study we examined the influence of chronic low-dose-rate irradiation on lifespan. Young male B6C3F1/Jcl mice were divided randomly into two groups, which were fed either a low-calorie (65 kcal/ week) or high-calorie (95 kcal/week) diet. The latter is comparable to ad libitum feeding. The animals in the irradiated group were continuously exposed to gamma rays for 400 days at 20 mGy/day, resulting in a total dose of 8 Gy. Exposure and calorie restriction were initiated at 8 weeks of age and the diets were maintained for life. The life-shortening effects from chronic whole-body irradiation were compared between the groups. Body weights were reduced in calorie-restricted mice irrespective of radiation treatment. Radiation induced a shortened median lifespan in both groups, but to a greater extent in the calorie-restricted mice. These results suggest that calorie restriction may sensitize mice to chronic low-dose-rate radiation exposure to produce a life-shortening effect rather than alleviating the effects of radiation.

Funding for radiation research: past, present and future.

For more than a century, ionizing radiation has been indispensable mainly in medicine and industry. Radiation research is a multidisciplinary field that investigates radiation effects. Radiation research was very active in the mid- to late 20th century, but has then faced challenges, during which time funding has fluctuated widely. Here we review historical changes in funding situations in the field of radiation research, particularly in Canada, European Union countries, Japan, South Korea, and the US. We also provide a brief overview of the current situations in education and training in this field. A better understanding of the biological consequences of radiation exposure is becoming more important with increasing public concerns on radiation risks and other radiation literacy. Continued funding for radiation research is needed, and education and training in this field are also important.

Sonographic sequential change of the anteroinferior labrum following arthroscopic Bankart repair: quantitative and qualitative assessment.

OBJECTIVE: Although arthroscopic Bankart repair is widely performed, little is known concerning the healing process of the Bankart lesion. This study aimed to describe the sonographic sequential change of the anteroinferior labrum following arthroscopic Bankart repair, both quantitatively and qualitatively. MATERIALS AND METHODS: Twenty-five subjects who had undergone arthroscopic Bankart repair were investigated using a sonographic diagnostic device equipped with shear-wave elastographic technology. In addition to measuring the elasticity of the anteroinferior labrum, the B-mode echogram of the anteroinferior labrum was classified into three grades based on the relative echogenicity. Assessment was performed in the affected shoulder 1, 2, 3, 4, 5, 6, and 12 months postoperatively and in the contralateral shoulder 1 month postoperatively for the control. RESULTS: The mean elasticity of the anteroinferior labrum in the affected shoulder 1 and 2 months postoperatively was significantly lower than in the contralateral shoulder (p < 0.001 for both). However, no significant difference was found after 3 months postoperatively. B-mode echograms of the anteroinferior labrum in the contralateral shoulder were classified as grade 0 in all subjects (100%), whereas the percentage of grade 0s in the affected shoulder was 0, 4, 96, and 100% at 1, 2, 3, and 4 months postoperatively. CONCLUSION: Both quantitative and qualitative assessment of repaired anteroinferior labrum using ultrasound became comparable with the contralateral shoulder 3-4 months postoperatively. Ultrasound is a useful decision-supporting tool to prescribe postoperative rehabilitation protocol following arthroscopic Bankart repair, although functional recovery should also be evaluated on an individual basis.

Experimental studies on the biological effects of chronic low dose-rate radiation exposure in mice: overview of the studies at the Institute for Environmental Sciences.

This review summarizes the results of experiments conducted in the Institute for Environmental Sciences for the past 21 years, focusing on the biological effects of long-term low dose-rate radiation exposure on mice. Mice were chronically exposed to gamma rays at dose-rates of 0.05, 1 or 20 mGy/day for 400 days to total doses of 20, 400 or 8000 mGy, respectively. The dose rate 0.05 mGy/day is comparable to the dose limit for radiation workers. The parameters examined were lifespan, neoplasm incidence, antineoplasm immunity, body weight, chromosome aberration(s), gene mutation(s), alterations in mRNA and protein levels and trans-generational effects. At 20 mGy/day, all biological endpoints were significantly altered except neoplasm incidence in the offspring of exposed males. Slight but statistically significant changes in lifespan, neoplasm incidences, chromosome abnormalities and gene expressions were observed at 1 mGy/day. Except for transient alterations in the mRNA levels of some genes and increased liver neoplasm incidence attributed to radiation exposure, the remaining biological endpoints were not influenced after exposure to 0.05 mGy/day. Results suggest that chronic low dose-rate exposure may induce small biological effects.

Differences in sustained alterations in protein expression between livers of mice exposed to high-dose-rate and low-dose-rate radiation.

Molecular mechanisms of radiation dose-rate effects are not well understood. Among many possibilities, long-lasting sustained alterations in protein levels would provide critical information. To evaluate sustained effects after acute and chronic radiation exposure, we analyzed alterations in protein expression in the livers of mice. Acute exposure consisted of a lethal dose of 8 Gy and a sublethal dose of 4 Gy, with analysis conducted 6 days and 3 months after irradiation, respectively. Chronic irradiation consisted of a total dose of 8 Gy delivered over 400 days (20 mGy/day). Analyses following chronic irradiation were done immediately and at 3 months after the end of the exposure. Based on antibody arrays of protein expression following both acute lethal and sublethal dose exposures, common alterations in the expression of two proteins were detected. In the sublethal dose exposure, the expression of additional proteins was altered 3 months after irradiation. Immunohistochemical analysis showed that the increase in one of the two commonly altered proteins, MyD88, was observed around blood vessels in the liver. The alterations in protein expression after chronic radiation exposure were different from those caused by acute radiation exposures. Alterations in the expression of proteins related to inflammation and apoptosis, such as caspase 12, were observed even at 3 months after the end of the chronic radiation exposure. The alterations in protein expression depended on the dose, the dose rate, and the passage of time after irradiation. These changes could be involved in long-term effects of radiation in the liver.

High Mutation Levels are Compatible with Normal Embryonic Development in Mlh1-Deficient Mice.

To elucidate the role of the mismatch repair gene Mlh1 in genome instability during the fetal stage, spontaneous mutations were studied in Mlh1-deficient lacZ-transgenic mouse fetuses. Mutation levels were high at 9.5 days post coitum (dpc) and gradually increased during the embryonic stage, after which they remained unchanged. In addition, mutations that were found in brain, liver, spleen, small intestine and thymus showed similar levels and no statistically significant difference was found. The molecular nature of mutations at 12.5 dpc in fetuses of Mlh1+/+ and Mlh1-/- mice showed their own unique spectra, suggesting that deletion mutations were the main causes in the deficiency of the Mlh1 gene. Of note, fetuses of irradiated mice exhibited marked differences such as post-implantation loss and Mendelian distribution. Collectively, these results strongly suggest that high mutation ofMlh1-/--deficient fetuses has little effect on the fetuses during their early developmental stages, whereas Mlh1-/--deficient fetuses from X-ray irradiated mothers are clearly effected.

Dose and dose-rate effects of ionizing radiation: a discussion in the light of radiological protection.

The biological effects on humans of low-dose and low-dose-rate exposures to ionizing radiation have always been of major interest. The most recent concept as suggested by the International Commission on Radiological Protection (ICRP) is to extrapolate existing epidemiological data at high doses and dose rates down to low doses and low dose rates relevant to radiological protection, using the so-called dose and dose-rate effectiveness factor (DDREF). The present paper summarizes what was presented and discussed by experts from ICRP and Japan at a dedicated workshop on this topic held in May 2015 in Kyoto, Japan. This paper describes the historical development of the DDREF concept in light of emerging scientific evidence on dose and dose-rate effects, summarizes the conclusions recently drawn by a number of international organizations (e.g., BEIR VII, ICRP, SSK, UNSCEAR, and WHO), mentions current scientific efforts to obtain more data on low-dose and low-dose-rate effects at molecular, cellular, animal and human levels, and discusses future options that could be useful to improve and optimize the DDREF concept for the purpose of radiological protection.

Skin can control solar UVR-induced mutations through the epidermis-specific response of mutation induction suppression.

Skin exposure to solar ultraviolet radiation (UVR) has been a major public concern because of its genotoxicity. We established recently three action spectra of UVR biological effects using inflammation, mutagenicity, and mutation induction suppression (MIS) as indicators to evaluate UVR risk for mammalian skin. MIS is an antigenotoxic epidermis-specific response by which the increase of the mutant frequency (MF) levels off above a certain UVR dose. Here, based on these spectra, the mutation load of the skin after sunlight exposure was evaluated utilizing the spectral solar-UVR intensity data which had been measured at Tsukuba, Japan by the Japan Meteorological Agency. We estimated the daily variation of the solar-UVR effectiveness (effect per second) for the three indicators, and revealed that the effectiveness efficiency (effect per dose) of midday sunlight is 3-4-fold higher than those in the early morning and late afternoon. Based on the daily variations of mutagenicity and MIS effectiveness, we further estimated MFs induced after every one-hour sunlight exposure and reached a remarkable prediction that MFs should be suppressed to a constant level during 9:00-15:00 by MIS. The estimates agreed well with the equivalent values directly determined at Sendai, a site close to Tsukuba, although a small difference was detected for the epidermis at the dose range where the suppressed MFs were predicted. We propose the use of observed minimum inflammation/erythema doses to improve the difference. Our method could provide reliable estimates of sunlight genotoxicity to evaluate skin cancer probabilities.

Solar-UV-signature mutation prefers TCG to CCG: extrapolative consideration from UVA1-induced mutation spectra in mouse skin.

UVA1 exerts its genotoxicity on mammalian skin by producing cyclobutane pyrimidine dimers (CPDs) in DNA and preferentially inducing solar-UV-signature mutations, C → T base substitution mutations at methylated CpG-associated dipyrimidine (Py-mCpG) sites, as demonstrated previously using a 364 nm laser as a UVA1 source and lacZ-transgenic mice that utilize the transgene as a mutational reporter. In the present study, we confirmed that a broadband UVA1 source induced the same mutation profiles in mouse epidermis as the UVA1 laser, generalizing the previous result from a single 364 nm to a wider wavelength range of UVA1 (340-400 nm). Combined with our previous data on the mutation spectra induced in mouse epidermis by UVB, UVA2 and solar UVR, we proved that the solar-UV-signature mutation is commonly observed in the wavelength range from UVB to UVA, and found that UVA1 induces this mutation more preferentially than the other shorter wavelength ranges. This finding indicates that the solar-UV-signature mutation-causing CPDs, which are known to prefer Py-mCpG sites, could be produced with the energy provided by the longer wavelength region of UVR, suggesting a photochemical reaction through the excitation of pyrimidine bases to energy states that can be accomplished by absorption of even low-energy UVR. On the other hand, the lower proportions of solar-UV-signature mutations observed in the mutation spectra for UVB and solar UVR indicate that the direct photochemical reaction through excited singlet state of pyrimidine bases, which can be accomplished only by high-energy UVR, is also involved in the mutation induction at those shorter wavelengths of UVR. We also found that the solar-UV signature prefers 5'-TCG-3' to 5'-CCG-3' as mutational target sites, consistent with the fact that UVA induces CPDs selectively at thymine-containing dipyrimidine sites and that solar UVR induces them preferably at Py-mCpG sites. However, the mutation spectrum in human p53 gene from non-melanoma skin cancers shows the opposite preference for 5'-CCG-3' sites. This apparent discrepancy in the site preference seems to result from the lack of 5'-TCG-3' sites mutable to missense mutations on the nontranscribed strand of human p53 gene, which should be evolutionally acquired under selective pressure from the sun.

Role of the Msh2 gene in genome maintenance and development in mouse fetuses.

In an attempt to evaluate the roles of the mismatch repair gene Msh2 in genome maintenance and in development during the fetal stage, spontaneous mutations and several developmental indices were studied in Msh2-deficient lacZ-transgenic mouse fetuses. Mutation levels in fetuses were elevated at 9.5 dpc (days post coitum) when compared to wild-type mice, and the level of mutations continued to increase until the fetuses reached the newborn stage. The mutation levels in 4 different tissues of newborns showed similar magnitudes to those in the whole body. The levels remained similar after birth until 6 months of age. The molecular nature of the mutations examined in 12.5 dpc fetuses of Msh2(+/+) and Msh2(-/-) revealed unique spectra which reflect errors produced during the DNA replication process, and those corrected by a mismatch repair system. Most base substitutions and simple deletions were reduced by the presence of the Msh2 gene, whereas G:C to A:T changes at CpG sequences were not affected, suggesting that the latter change was not influenced by mismatch repair. On the other hand, analysis of developmental indices revealed that there was very little effect, including the presence of malformations, resulting from Msh2-deficiencies. These results indicate that elevated mutation levels have little effect on the development of the fetus, even if a mutator phenotype appears at the organogenesis stage.

Fully functional global genome repair of (6-4) photoproducts and compromised transcription-coupled repair of cyclobutane pyrimidine dimers in condensed mitotic chromatin.

During mitosis, chromatin is highly condensed, and activities such as transcription and semiconservative replication do not occur. Consequently, the condensed condition of mitotic chromatin is assumed to inhibit DNA metabolism by impeding the access of DNA-transacting proteins. However, about 40 years ago, several researchers observed unscheduled DNA synthesis in UV-irradiated mitotic chromosomes, suggesting the presence of excision repair. We re-examined this subject by directly measuring the removal of UV-induced DNA lesions by an ELISA and by a Southern-based technique in HeLa cells arrested at mitosis. We observed that the removal of (6-4) photoproducts from the overall genome in mitotic cells was as efficient as in interphase cells. This suggests that global genome repair of (6-4) photoproducts is fully functional during mitosis, and that the DNA in mitotic chromatin is accessible to proteins involved in this mode of DNA repair. Nevertheless, not all modes of DNA repair seem fully functional during mitosis. We also observed that the removal of cyclobutane pyrimidine dimers from the dihydrofolate reductase and c-MYC genes in mitotic cells was very slow. This suggests that transcription-coupled repair of cyclobutane pyrimidine dimers is compromised or non-functional during mitosis, which is probably the consequence of mitotic transcriptional repression.

A case of mediastinal lymph node carcinoma of unknown primary site treated with docetaxel and cisplatin with concurrent thoracic radiation therapy.

Mediastinal lymph node carcinoma of unknown primary site is rare and may have a better prognosis if extensive treatment is performed. Case, A 69-year-old-male presented with a persistent cough. Chest computed tomography (CT) demonstrated a large tumor 9.5 × 8.2 cm, in the mediastinum, compressing the right main bronchus, the right pulmonary artery, and the superior vena cava. Because fiberoptic bronchoscopy was insufficient for diagnosis, mediastinoscopic tumor biopsy under general anesthesia was undertaken. Histological examination revealed adenocarcinoma. Extensive examinations revealed no other neoplastic lesion except in the mediastinum. Mediastinal lymph node carcinoma of unknown primary site was diagnosed. The patient was treated with docetaxel and cisplatin with concurrent thoracic radiation therapy. A month after the start of chemoradiotherapy, the mediastinal tumor regressed markedly. The patient remained free of symptoms without regrowth of the primary site. Exploration of the body showed no further abnormalities 20 months after disease onset.

Antigenotoxic effects of p53 on spontaneous and ultraviolet light B--induced deletions in the epidermis of gpt delta transgenic mice.

Tumor development in the skin may be a multistep process where multiple genetic alterations occur successively. The p53 gene is involved in genome stability and thus is referred to as "the guardian of the genome." To better understand the antigenotoxic effects of p53 in ultraviolet light B (UVB)-induced mutagenesis, mutations were measured in the epidermis of UVB-irradiated p53(+/+) and p53(-/-) gpt delta mice. In the mouse model, point mutations and deletions are separately identified by the gpt and Spi(-) assays, respectively. The mice were exposed to UVB at single doses of 0.5, 1.0, or 2.0 kJ/m(2) . The mutant frequencies (MFs) were determined 4 weeks after the irradiation. All doses of UVB irradiation enhanced gpt MFs by about 10 times than that of unirradiated mice. There were no significant differences in gpt MFs and the mutation spectra between p53(+/+) and p53(-/-) mice. The predominant mutations induced by UVB irradiation were G:C to A:T transitions at dipyrimidines. In contrast, in unirradiated p53(-/-) mice, the frequencies of Spi(-) large deletions of more than 1 kb and complex-type deletions with rearrangements were significantly higher than those of the Spi(-) large deletions in p53(+/+) counterparts. The specific Spi(-) mutation frequency of more than 1 kb deletions and complex types increased in a dose-dependent manner in the p53(+/+) mice. However, no increase of such large deletions was observed in irradiated p53(-/-) mice. These results suggest that the antigenotoxic effects of p53 may be specific to deletions and complex-type mutations induced by double-strand breaks in DNA.

Gene expression profiles in mouse liver after long-term low-dose-rate irradiation with gamma rays.

Changes in gene expression profiles in mouse liver induced by long-term low-dose-rate γ irradiation were examined by microarray analysis. Three groups of male C57BL/6J mice were exposed to whole-body radiation at dose rates of 17-20 mGy/day, 0.86-1.0 mGy/day or 0.042-0.050 mGy/day for 401-485 days with cumulative doses of approximately 8 Gy, 0.4 Gy or 0.02 Gy, respectively. The gene expression levels in the livers of six animals from each exposure group were compared individually with that of pooled sham-irradiated animals. Some genes revealed a large variation in expression levels among individuals within each group, and the number of genes showing common changes in individuals from each group was limited: 20 and 11 genes showed more than 1.5-fold modulation with 17-20 mGy/day and 0.86-1.0 mGy/day, respectively. Three genes showed more than 1.5-fold modulation even at the lowest dose-rate of 0.04-0.05 mGy/day. Most of these genes were down-regulated. RT-PCR analysis confirmed the expression profiles of the majority of these genes. The results indicate that a few genes are modulated in response to very low-dose-rate irradiation. The functional analysis suggests that these genes may influence many processes, including obesity and tumorigenesis.

Influences of p53 deficiency on the apoptotic response, DNA damage removal and mutagenesis in UVB-exposed mouse skin.

p53 suppresses the genomic instability provoked by genotoxic agents. Ultraviolet (UV) B induces skin cancers by producing DNA damage and mutations in the skin genome, whereas the skin tissue responds to the UVB insult with cell cycle arrest and apoptosis as well as damage exclusion by DNA repair. To address the p53 contribution to these skin responses in vivo, we analyzed the time course of DNA damage removal, apoptosis induction and hyperplasia in the skin after UVB irradiation in p53-knockout mice. We also examined UVB-induced mutations in the skin. We found that p53 deficiency does not abolish the UVB-induced apoptotic response in the epidermis but delays the process and the following hyperplasia 12-24 h. Regardless of the p53 genotype, 1 kJ/m(2) UVB induced a total replacement of the epidermal layer by destroying the damaged epidermis by apoptosis and rebuilding a new one through hyperplasia. We failed to detect a clear defect in removal of UVB-induced DNA photolesions from the genome of the p53-deficient skin except for a delay in the epidermis, which seemed to result from the delay in the apoptotic response. However, we found that p53 deficiency enhanced UVB-induced mutagenesis. Furthermore, in a genetic study using Xpa-knockout mice, we showed that the enhanced mutagenic response depends on the activity of nucleotide excision repair (NER), which was also supported by the mutation spectrum observed in the UVB-exposed p53-knockout mice. These results indicate that p53 protects the skin genome from the UVB genotoxicity by facilitating NER, whereas its contribution to the UVB-induced apoptosis is limited.

Radiation dose-rate effect on mutation induction in spleen and liver of gpt delta mice.

The effect of dose rate on radiation-induced mutations in two somatic tissues, the spleen and liver, was examined in transgenic gpt delta mice. These mice can be used for the detection of deletion-type mutations, and these are the major type of mutation induced by radiation. The dose rates examined were 920 mGy/min, 1 mGy/min and 12.5 microGy/min. In both tissues, the number of mutations increased with increasing dose at each of the three dose rates examined. The mutation induction rate was dependent on the dose rate. The mutation induction rate was higher in the spleen than in the liver at the medium dose rate but was similar in the two tissues at the high and low dose rates. The mutation induction rate in the liver did not show much change between the medium and low dose rates. Analysis of the molecular nature of the mutations indicated that 2- to 1,000-bp deletion mutations were specifically induced by radiation in both tissues after high- and low-dose-rate irradiation. The occurrence of deletion mutation without any sequence homology at the break point was elevated in spleen after high-dose-rate irradiation. The results indicate that the mutagenic effects of radiation in somatic tissues are dependent on dose rate and that there is some variability between tissues.

XPC is involved in genome maintenance through multiple pathways in different tissues.

In an attempt to evaluate the role of the Xpc gene in maintaining genomic stability in vivo under normal conditions, the age-dependent accumulation of spontaneous mutations in different tissues was analyzed in Xpc-deficient lacZ-transgenic mice. Brain, testis, and small intestine revealed no effects from the Xpc-deficiency, whereas liver, spleen, heart, and lung showed an enhanced age-related accumulation of mutations in Xpc-deficient mice. In the spleen, the effect was not obvious at 2 and 12 months of age, but became apparent at 23 months. The magnitude of the observed effect at an advanced age was similar in the liver, spleen and heart, but was comparatively smaller in the lung. Haploinsufficiency was observed in liver and spleen but not in heart and lung. Analysis of DNA sequences in the mutants revealed that the frequency of G:C to T:A changes were elevated in the liver and heart of Xpc-deficient aged mice, supporting the possible involvement of XPC in base excision repair of oxidized guanine. The occurrence of two or more mutations within a single lacZ gene was termed a multiple mutation and was also elevated in old Xpc-deficient mice. Among the clones examined, two mutant clones showed as many as four mutations within a short stretch of DNA. This is the first demonstration to support suggestions for the existence of a role for XPC in the suppression of multiple mutations. These multiple mutations could conceivably be generated by error-prone trans-lesional DNA synthesis. Overall, these results indicate that there may be diverse roles or mechanisms through which XPC participates in genome maintenance in different tissues.

Induction of mitosis delay, apoptosis and aneuploidy in human cells by phenyl hydroquinone, an Ames test-negative carcinogen.

Ortho-phenyl phenol and its hepatic derivative, phenyl hydroquinone, do not generate base-substitution-type mutations, but cause bladder cancer in rats and mice. The mechanism of their carcinogenic effect is unknown. We have previously shown that o-phenyl phenol and phenyl hydroquinone induce mitotic arrest and aneuploidy in Saccharomyces cerevisiae. To further delineate the mechanism of action of phenyl hydroquinone, we examined its effect on human cells. Treatment of the colon cancer cell line HCT116 with 0 to 150 microM phenyl hydroquinone caused a concentration-dependent inhibition of growth, accumulation of cells having G2/M DNA content, and an increase in the mitotic index. Moreover, a dose-dependent increase in apoptotic cells was observed. Finally, a high frequency of aneuploid cells was found. On the other hand, no increase in gamma-H2AX foci was observed. The results show that phenyl hydroquinone does induce mitotic arrest, apoptosis and aneuploidy in the absence of DNA damage. Our results may be useful to understand the mechanisms of action of chemical substances that are Ames test-negative carcinogens.

Functional interactions between BLM and XRCC3 in the cell.

Bloom's syndrome (BS), which is caused by mutations in the BLM gene, is characterized by a predisposition to a wide variety of cancers. BS cells exhibit elevated frequencies of sister chromatid exchanges (SCEs), interchanges between homologous chromosomes (mitotic chiasmata), and sensitivity to several DNA-damaging agents. To address the mechanism that confers these phenotypes in BS cells, we characterize a series of double and triple mutants with mutations in BLM and in other genes involved in repair pathways. We found that XRCC3 activity generates substrates that cause the elevated SCE in blm cells and that BLM with DNA topoisomerase IIIalpha suppresses the formation of SCE. In addition, XRCC3 activity also generates the ultraviolet (UV)- and methyl methanesulfonate (MMS)-induced mitotic chiasmata. Moreover, disruption of XRCC3 suppresses MMS and UV sensitivity and the MMS- and UV-induced chromosomal aberrations of blm cells, indicating that BLM acts downstream of XRCC3.