Nondisjunction Research Articles

Light-induced blockage of cell division with a chromatin-targeted phototoxic fluorescent protein

Proteins of the GFP (green fluorescent protein) family are widely used as passive reporters for live cell imaging. In the present study we used H2B (histone H2B)-tKR (tandem KillerRed) as an active tool to affect cell division with light. We demonstrated that H2B-tKR-expressing cells behave normally in the dark, but transiently cease proliferation following green-light illumination. Complete light-induced blockage of cell division for approx. 24 h was observed in cultured mammalian cells that were either transiently or stably transfected with H2B-tKR. Illuminated cells then returned to normal division rate. XRCC1 (X-ray cross complementing factor 1) showed immediate redistribution in the illuminated nuclei of H2B-tKR-expressing cells, indicating massive light-induced damage of genomic DNA. Notably, nondisjunction of chromosomes was observed for cells that were illuminated during metaphase. In transgenic Xenopus embryos expressing H2B-tKR under the control of tissue-specific promoters, we observed clear retardation of the development of these tissues in green-light-illuminated tadpoles. We believe that H2B-tKR represents a novel optogenetic tool, which can be used to study mitosis and meiosis progression per se, as well as to investigate the roles of specific cell populations in development, regeneration and carcinogenesis in vivo.

Terahertz Radiation Induces Spindle Disturbances in Human-Hamster Hybrid Cells

The aim of this study was to investigate and quantify the production of spindle disturbances in A(L) cells, a human-hamster hybrid cell line, by 0.106 THz radiation (continuous wave). Monolayer cultures in petri dishes were exposed for 0.5 h to 0.106 THz radiation with power densities ranging from 0.043 mW/cm(2) to 4.3 mW/cm(2) or were kept under sham conditions (negative control) for the same period. As a positive control, 100 µg/ml of the insecticide trichlorfon, which is an aneuploidy-inducing agent, was used for an exposure period of 6 h. During exposure, the sample containers were kept at defined environmental conditions in a modified incubator as required by the cells. Based on a total of 6,365 analyzed mitotic cells, the results of two replicate experiments suggest that 0.106 THz radiation is a spindle-acting agent as predominately indicated by the appearance of spindle disturbances at the anaphase and telophase (especially lagging and non-disjunction of single chromosomes) of cell divisions. The findings in the present study do not necessarily imply disease or injury but may be important for evaluating possible underlying mechanisms.

Chromosome Abnormalities in the Human Oocyte

Aneuploidy is the most commonly occurring type of chromosome abnormality and the most significant clinically. It arises mostly due to segregation errors taking place during female meiosis and is also closely associated with advancing maternal age. Two main aneuploidy-causing mechanisms have been described: the first involves the non-disjunction of entire chromosomes and can take place during both meiotic divisions, whereas the second involves the premature division of a chromosome into its 2 sister chromatids, followed by their random segregation, upon completion of meiosis I. To elucidate the causal mechanisms of maternally derived aneuploidy and the manner with which they affect the 2 meiotic divisions, a large number of oocytes and their corresponding polar bodies have been examined. Various classical and molecular cytogenetic methods have been employed for this purpose, and valuable data have been obtained. Moreover, research into the gene expression patterns of oocytes according to maturity, maternal age, and chromosome status has provided a unique insight into the complex nature of the biological processes and genetic pathways regulating female meiosis. Findings obtained from the cytogenetic and molecular analysis of oocytes will be reviewed in this article.

Chromosomal and cytoplasmic context determines predisposition to maternal age-related aneuploidy: brief overview and update on MCAK in mammalian oocytes

It has been known for more than half a century that the risk of conceiving a child with trisomy increases with advanced maternal age. However, the origin of the high susceptibility to nondisjunction of whole chromosomes and precocious separation of sister chromatids, leading to aneuploidy in aged oocytes and embryos derived from them, cannot be traced back to a single disturbance and mechanism. Instead, analysis of recombination patterns of meiotic chromosomes of spread oocytes from embryonal ovary, and of origins and exchange patterns of extra chromosomes in trisomies, as well as morphological and molecular studies of oocytes and somatic cells from young and aged females, show chromosome-specific risk patterns and cellular aberrations related to the chronological age of the female. In addition, analysis of the function of meiotic- and cell-cycle-regulating genes in oogenesis, and the study of the spindle and chromosomal status of maturing oocytes, suggest that several events contribute synergistically to errors in chromosome segregation in aged oocytes in a chromosome-specific fashion. For instance, loss of cohesion may differentially predispose chromosomes with distal or pericentromeric chiasmata to nondisjunction. Studies on expression in young and aged oocytes from human or model organisms, like the mouse, indicate that the presence and functionality/activity of gene products involved in cell-cycle regulation, spindle formation and organelle integrity may be altered in aged oocytes, thus contributing to a high risk of error in chromosome segregation in meiosis I and II. Genes that are often altered in aged mouse oocytes include MCAK (mitotic-centromere-associated protein), a microtubule depolymerase, and AURKB (Aurora kinase B), a protein of the chromosomal passenger complex that has many targets and can also phosphorylate and regulate MCAK localization and activity. Therefore we explored the role of MCAK in maturing mouse oocytes by immunofluorescence, overexpression of a MCAK-EGFP (enhanced green fluorescent protein) fusion protein, knockdown of MCAK by RNAi (RNA interference) and inhibition of AURKB. The observations suggest that MCAK is involved in spindle regulation, chromosome congression and cell-cycle control, and that reductions in mRNA and protein in a context of permissive SAC (spindle assembly checkpoint) predispose to aneuploidy. Failure to recruit MCAK to centromeres and low expression patterns, as well as disturbances in regulation of enzyme localization and activity, e.g. due to alterations in activity of AURKB, may therefore contribute to maternal age-related rises in aneuploidy in mammalian oocytes.

Structural chromosomal aberrations, aneuploidy, and mosaicism in early cleavage mouse embryos derived from spermatozoa exposed to γ-rays

Purpose: To quantitatively and qualitatively investigate the changes in chromosomal aberrations during early cleavage in mouse embryos derived from γ-irradiated spermatozoa.Materials and methods: Mature males were exposed to 2 Gy or 4 Gy of 137Cs γ-rays, and their spermatozoa were used to produce embryos via in vitro fertilisation (IVF). The metaphase chromosomes were prepared from one-cell, two-cell, and four-cell embryos. In the chromosome preparations from two-cell and four-cell embryos, the separation of the sister blastomeres was precluded by treatment of the embryos with concanavalin A. The incidence of embryos with structural chromosomal aberrations, aneuploidy, or mosaicism was estimated. The fates of the different types of γ-ray-induced structural chromosomal aberrations were also investigated in those embryos.Results: The exposure of spermatozoa to 2 Gy or 4 Gy γ-rays caused structural chromosomal aberrations in 25.9% and 35.7% of the resultant one-cell embryos, respectively. At two-cell embryonic stage, the incidence of structural chromosomal aberrations was 17.4% in the 2 Gy group and 27.1% in the 4 Gy group. At the four-cell embryonic stage, although the incidence of control embryos with structural chromosomal aberrations was considerably high, the net incidence of embryos with radiation-induced structural chromosomal aberrations was similar to that at the one-cell stage. The incidence of aneuploidy was high in two-cell and four-cell embryos after both doses of γ-rays. The incidence of mosaicism increased significantly in dose- and embryonic-stage-dependent manners. Anaphase lag, and the degeneration and non-disjunction of the aberrant chromosomes were frequently observed in aneuploid and mosaic embryos.Conclusions: Mouse sperm DNA is highly vulnerable to γ-rays. The structural chromosomal aberrations of sperm origin are unstable in their behaviour and structure during cleavage, and therefore cause secondary aneuploidy and mosaicism in the early cleavage embryos.

Effect of vitrification and thawing on human oocyte ATP concentration

Vitrification/thawing has a significant negative impact on oocyte ATP concentration. However, incubating oocytes up to 180 minutes after thawing allowed oocytes to regain some ATP level.

Cytogenetic mechanisms of aneuploidy in somatic cells of chemonuclear industry professionals with incorporated plutonium-239

Risks related to occupational exposure of humans to ionizing radiation remains a focus of attention. Of special importance is the poorly understood effect of ionizing radiation on the rate of numerical chromosome aberrations in human somatic cells. The purpose of this study was characterization of a radiation-induced increase in the rate of aneuploidy induced by incorporated plutonium-239 (239Pu) in somatic cells of chemonuclear industry workers. Significant differences in the chromosome 2, 7, 8, 12, and Y aneuploidy rate have been found between subjects with incorporated 239Pu and control subjects have been demonstrated. Chromosome nondisjunction has been found to be the main mechanism of aneuploidy induced by incorporated 239Pu.

Statistical Analysis of Nondisjunction Assays in Drosophila

Many advances in the understanding of meiosis have been made by measuring how often errors in chromosome segregation occur. This process of nondisjunction can be studied by counting experimental progeny, but direct measurement of nondisjunction rates is complicated by not all classes of nondisjunctional progeny being viable. For X chromosome nondisjunction in Drosophila female meiosis, all of the normal progeny survive, while nondisjunctional eggs produce viable progeny only if fertilized by sperm that carry the appropriate sex chromosome. The rate of nondisjunction has traditionally been estimated by assuming a binomial process and doubling the number of observed nondisjunctional progeny, to account for the inviable classes. However, the correct way to derive statistics (such as confidence intervals or hypothesis testing) by this approach is far from clear. Instead, we use the multinomial-Poisson hierarchy model and demonstrate that the old estimator is in fact the maximum-likelihood estimator (MLE). Under more general assumptions, we derive asymptotic normality of this estimator and construct confidence interval and hypothesis testing formulae. Confidence intervals under this framework are always larger than under the binomial framework, and application to published data shows that use of the multinomial approach can avoid an apparent type 1 error made by use of the binomial assumption. The current study provides guidance for researchers designing genetic experiments on nondisjunction and improves several methods for the analysis of genetic data.

Xnd-1 regulates the global recombination landscape in Caenorhabditis elegans

Meiotic crossover (CO) recombination establishes physical linkages between homologous chromosomes that are required for their proper segregation into developing gametes and promotes genetic diversity by shuffling genetic material between parental chromosomes. COs require the formation of double strand breaks (DSBs) to create the substrate for strand exchange. DSBs occur in small intervals called hotspots1-3 and significant variation in hotspot usage exists between and among individuals4. This variation is thought to reflect differences in sequence identity and chromatin structure, DNA topology and/ or chromosome domain organization1, 5-9. Chromosomes show different frequencies of nondisjunction (NDJ)10, reflecting inherent differences in meiotic crossover control, yet the underlying basis of these differences remains elusive. Here we show that a novel chromatin factor, X non-disjunction factor 1 (xnd-1), is responsible for the global distribution of COs in C. elegans. xnd-1 is also required for formation of double-strand breaks (DSBs) on the X, but surprisingly XND-1 protein is autosomally-enriched. We show that xnd-1 functions independently of genes required for X chromosome-specific gene silencing, revealing a novel pathway that distinguishes the X from autosomes in the germ line, and further show that xnd-1 exerts its effects on COs, at least in part, by modulating levels of H2A lysine 5 acetylation.

Comprehensive embryo analysis of advanced maternal age–related aneuploidies and mosaicism by short comparative genomic hybridization

The short comparative genomic hybridization (short-CGH) method was used to perform a comprehensive cytogenetic study of isolated blastomeres from advanced maternal age embryos, discarded after fluorescent in situ hybridization (FISH) preimplantation genetic screening (PGS), detecting aneuploidies (38.5% of which corresponded to chromosomes not screened by 9-chromosome FISH), structural aberrations (31.8%), and mosaicism (77.3%). The short-CGH method was subsequently applied in one PGS, achieving a twin pregnancy.

Chromosomal basis of evolution in the genus Coix L. (Maydeae): a critical appraisal

Among the nine species of the genus Coix, C. lacrymajobi L. and C. gigantea Koen. have 2n = 20 small chromosomes and C. aquatica Roxb. 2n = 10 large chromosomes, and are widely distributed in South and South East Asia, the first is cultivated and spread to all warmer parts of the world. The other species have restricted localized distribution. C. aquatica originated from a hypothetical diploid ancestor with 2n = 10 small chromosomes, which became longer through accumulation of duplicate segments and heterochromatin and adapted to wet habitats. C. lacrymajobi and C. gigantea originated through the same two small chromosome diploid ancestors, one of which may be common to C. aquatica, through allopolyploidy and adapted to drier conditions. All the three species can survive equally well in wet and dry localities. Aneuploid and polyploid races occurred through chromosome nondisjunction and genome doubling in C. gigantea and C. aquatica, generation of alien-addition and substitution chromosome races appears to be the further course of evolution. Six morphological varieties were differentiated in C. lacrymajobi through accumulation of gene mutations, and chromosome structural changes gave rise to a completely euchromatic chromosome race. Two aneuploid species with 2n = 32 chromosomes, both large and small, must have originated through allopolyploidy, with C. aquatica (n = 6) race as one of the parents in both, C. lacrymajobi and C. gigantea (n = 10) the second parent in Madhya Pradesh form and West Bengal form respectively. From comparison of morphology and their distribution, C. puellarum Balansa, C. ouwehandii Koord. and C. poilanei Mimeur are speculated to have originated from the three established species, through chromosomal changes and gene mutations and adapted to restricted localized areas, and C. gasteenii Simon an allopolyploid form of C. lacrymajobi and C. gigantea. A schematic diagram is presented depicting the origin, evolution and interrelationships in the genus.

Spermatogenesis in house mouse in a Robertsonian polymorphism zone

Robertsonian (Rb) translocations can be important in speciation as a mechanism of postzygotic isolation between populations. Meiotic non-disjunction, gametogenic impairment, and association of impaired autosomal segments with sex chromosomes have been postulated as mechanisms responsible for reducing fertility in Rb mice. Quantitative histological studies needed to understand the role of Rb fusions in gametogenic impairment are scarce. Most research on Rb mice has analyzed meiotic non-disjunction of laboratory and wild-derived strains, which have complex or simple structural heterozygosity with large numbers of fusions. Using histological multilevel sampling, we examined spermatogenesis in mice from the Rb polymorphism area of Barcelona. We studied four chromosomal groups having: a) one Rb heterozygote fusion and 2n=39, b) one Rb heterozygote fusion and 2n=31, c) three Rb heterozygote fusions without monobrachial homology and with diploid number ranging from 2n=29 to 2n=37, and d) only Rb homozygote fusions with diploid number ranging from 2n=28 to 2n=30. Standard mice from the area surrounding the Rb zone were used as control. We analyzed morphological variables of the testes, relative frequency of stages in the seminiferous epithelium cycle, the 'round spermatids:primary spermatocytes' ratio, and other derived parameters. Our results reveal that structural homozygote mice and simple heterozygote mice having as few as one to three Rb fusions undergo greater germ cell death (GCD) than standard mice, suggesting that Rb fusions are related to increased GCD (in both the heterozygous and homozygous state) and may be the main cause of decreased gene flow between mice populations from this area.

Human Embryonic Stem Cells as Models for Aneuploid Chromosomal Syndromes

Syndromes caused by chromosomal aneuploidies are widely recognized genetic disorders in humans and often lead to spontaneous miscarriage. Preimplantation genetic screening is used to detect chromosomal aneuploidies in early embryos. Our aim was to derive aneuploid human embryonic stem cell (hESC) lines that may serve as models for human syndromes caused by aneuploidies. We have established 25 hESC lines from blastocysts diagnosed as aneuploid on day 3 of their in vitro development. The hESC lines exhibited morphology and expressed markers typical of hESCs. They demonstrated long-term proliferation capacity and pluripotent differentiation. Karyotype analysis revealed that two-third of the cell lines carry a normal euploid karyotype, while one-third remained aneuploid throughout the derivation, resulting in eight hESC lines carrying either trisomy 13 (Patau syndrome), 16, 17, 21 (Down syndrome), X (Triple X syndrome), or monosomy X (Turner syndrome). On the basis of the level of single nucleotide polymorphism heterozygosity in the aneuploid chromosomes, we determined whether the aneuploidy originated from meiotic or mitotic chromosomal nondisjunction. Gene expression profiles of the trisomic cell lines suggested that all three chromosomes are actively transcribed. Our analysis allowed us to determine which tissues are most affected by the presence of a third copy of either chromosome 13, 16, 17 or 21 and highlighted the effects of trisomies on embryonic development. The results presented here suggest that aneuploid embryos can serve as an alternative source for either normal euploid or aneuploid hESC lines, which represent an invaluable tool to study developmental aspects of chromosomal abnormalities in humans.

Abnormal Villous Morphology Associated with Triple Trisomy of Paternal Origin

The vast majority of trisomies in spontaneous abortions (SAB) are single and of maternal origin, most frequently due to meiosis I errors. Triple trisomies are exceedingly rare (approximately 0.05% of spontaneous abortions), most often of maternal origin, and associated with increased maternal age. Some trisomic SAB specimens can exhibit abnormal villous morphology simulating a partial hydatidiform mole, a distinct form of hydatidiform mole characterized by diandric triploidy. A SAB specimen from a 27-year-old woman, G1P0 at 8 weeks gestational age, was reviewed in consultation to address the finding of morphological features suggestive of a partial hydatidiform mole but DNA ploidy analysis yielding a diploid result. The villi were irregularly shaped and hydropic but lacked trophoblastic hyperplasia; p57 expression was retained. Since fully developed features of a partial hydatidiform mole were lacking, additional analysis was performed. Molecular genotyping and single nucleotide polymorphism array analysis demonstrated biparental diploidy with trisomy of chromosomes 7, 13, and 20, all of paternal origin. The three trisomies may have originated from paternal meiosis II errors, or from mitotic nondisjunction. We believe this to be the first report of triple trisomy in a SAB confirmed to be of paternal origin.

Chromosome 21 non-disjunction and Down syndrome birth in an Indian cohort: analysis of incidence and aetiology from family linkage data

We analysed the family linkage data obtained from short tandem repeat (STR) genotyping of 212 unrelated Indian families having a single Down syndrome (DS) baby each, in order to explore the incidence and aetiology of this human aneuploidy in our cohort. The estimated values of maternal meiotic I and meiotic II non-disjunction (NDJ) errors of chromosome 21 (Ch 21) were approximately 78 and approximately 22%, respectively. Within the paternal outcome group, about 47 and 53% were accounted for NDJ at meiosis I and meiosis II, respectively. We estimated only approximately 2% post-zygotic mitotic errors. The comparison of average age of conception between controls and DS-bearing mothers revealed a significant difference (P<0.001) with DS-bearing women were on an average older than controls and meiotic II non-disjoined mothers were oldest among meiotic outcome groups. Our linkage analysis suggested an overall reduction in recombination by more than 50% on meiotic I non-disjoined maternal Ch 21 with error prone to susceptible chiasma formation within the approximately 5.1 kbp segment near the telomeric end. We stratified meiotic I non-disjoined women in three age groups, viz. young (<or=28 years), middle (29-34 years) and old (>or=35 years) and found linear decrease in the frequency of achiasmate meiosis from the young to the old group. In contrary, a linear increase in the multiple chiasma frequency from the young to the old group was observed. Considering these results together, we propose that the risk factors for Ch 21 NDJ are of two types, one being 'maternal age-independent' and the other being 'maternal age-dependent'. Moreover, a comparison of our present Indian dataset with that of other published data of ethnically different populations suggested that the genetics that underlies the NDJ of Ch 21 is probably universal irrespective of racial difference across human populations. The present study is the first population-based report on any DS cohort from the Indian subcontinent and our work will help future workers in understanding better the aetiology of this birth defect.

Androgenetic complete mole with trisomy 13: Report of a case with microsatellite genotyping and review of the literature

Hydatidiform moles are gestational diseases with abnormal development of the villous trophoblast and characterized by an excess of paternal to maternal genetic material. Complete moles are usually diploid and androgenetic, and are thought to develop after the fertilization of an “empty ovum” by either a haploid spermatozoon or two spermatozoa. We report a case of a complete mole in which fluorescence in situ hybridization (FISH) incidentally disclosed trisomy 13. Microsatellite genotyping showed a single allele at each of the markers tested on the chorionic villi, and comparison with parental peripheral blood specimens revealed that the markers were all of paternal origin. These results confirmed the paternal origin of all three copies of chromosome 13, and the isodisomy for each chromosome was consistent with duplication of a monospermic fertilization event and subsequent non-disjunction. To the best of our knowledge, this is the only case of an androgenetic complete mole with trisomy 13 described in the scientific literature. We present a review of the literature and hypothesize that the trisomy 13 in our case likely resulted from non-disjunction of chromosome 13.

A Dual Activity of ROS and Oxidative Stress on Adult Neurogenesis and Alzheimers Disease

Oxidative stress is a deleterious condition leading to cellular death. It plays a key role in the development and pathology of neurodegenerative diseases, like Alzheimer's disease (AD). AD is the most common form of dementia among elderly. Genetic mutations and genetic, acquired and environmental risk factors, particularly neuroinflammation and oxidative stress, are the main causes of AD. Neurogenesis occurs in the adult brain of mammals, particularly in the hippocampus, and is enhanced in the brain of patients with AD. Enhanced neurogenesis in AD may represent an attempt by the central nervous system to compensate for the neuronal loss and repair itself. Reactive oxygen species (ROS) promote cell death and the nondisjunction of chromosomes, leading to aneuploidy. The activity of ROS on newly generated neuronal cells in the adult brain may contribute to the pathogenesis of AD. Antioxidant may be used to reduce the deleterious activity of ROS, particularly on newly generated neuronal cells of the adult brain, potentially delaying the development of AD and promoting the regenerative capacity of the adult brain.

Trisomie 21 : 50 ans entre médecine et science

Fifty years after the discovery of the etiology of Down syndrome, trisomy 21 remains the model of choice for studying human diseases resulting from the presence of a chromosome or a chromosome segment in excess. In this review, mechanisms of aneuploidy occurrence and consequences of genomic imbalances will be mainly discussed. The study of genetic markers showed that trisomy 21 results in 90% of cases from an error during maternal meiosis. Approximately 8% of cases result from an error during paternal meiosis and in 2% of cases there is a postzygotic mitotic nondisjunction. The biological basis of the effect of maternal age remains largely unknown. The absence of genetic recombination between homologous chromosomes or the presence of an exchange in telomeric position are two risk factors of non-disjunction observed in young women. Non-disjunctions associated with pericentromeric exchanges are observed with an increase in maternal age. The study of mouse models and patients with partial trisomy 21, combined with advances in knowledge of the physical map and the transcriptome, identified genes directly or indirectly involved in the pathogenesis of Down syndrome. The recent description of metabolic pathways controlled by RCAN1 and DYRK1A genes which may be involved in many biological processes and phenotypes associated with trisomy 21 allows to consider new therapeutic strategies.

Genomic responses to abnormal gene dosage: the X chromosome improved on a common strategy.

Mechanisms to guard genomic integrity are critical to ensuring the welfare and survival of an organism. Disruptions of genomic integrity can result in aneuploidy, a large-scale genomic imbalance caused by either extra or missing whole chromosomes (chromosomal aneuploidy) or chromosome segments (segmental aneuploidy). A change in dosage of a single gene may not compromise the well-being of an organism, but the combined altered dosage of many genes due to aneuploidy disturbs the overall balance of gene expression networks, resulting in decreased fitness and mortality [1],[2]. Chromosomal aneuploidy is a common cause of birth defects—Down syndrome is caused by an extra copy of Chromosome 21, and Turner syndrome by a single copy of the X chromosome in females. Furthermore, methods that detect segmental aneuploidy have uncovered small deletions or duplications of the genome in association with many disorders, such as mental retardation. Chromosomal and segmental aneuploidies are also frequent in cancer cells in which changes in copy number paradoxically increase cell fitness but are unfavorable to survival of the organism. A fundamental issue in biology and medicine is to understand the effects of aneuploidy on gene expression and the mechanisms that alleviate aneuploidy-induced imbalance of the genome. Chromosomal aneuploidy is caused by non-disjunction of chromosomes in meiosis or mitosis, while segmental aneuploidy involves breakage and ligation of DNA. In contrast, the sex chromosomes provide an example of a naturally occurring “aneuploidy” caused by the evolution of a specific set of chromosomes for sex determination that often differ in their copy number between males and females. For example, in mammals and in flies, females have two X chromosomes and males have one X chromosome and a Y chromosome, resulting in X monosomy in males. How does a cell or an organism respond to such different types of aneuploidy, abnormal or natural? It turns out that the overall expression level of a given gene is not necessarily in direct relation to the copy number. Unique strategies have evolved to deal with abnormal gene dosage to alleviate the effects of aneuploidy by dampening changes in expression levels. What's more, the X chromosome has evolved sophisticated mechanisms to achieve complete dosage compensation, not surprisingly, since the copy number difference between males and females has been evolving for a long time.

Micronucleus-centromere assay in workers occupationally exposed to low level of benzene

Benzene is a well-known carcinogen that induces chromosomal instability, including chromosome aberration and aneuploidy. In order to assess aneugenic effect of low-level benzene, micronucleus-centromere assay using specific probes for chromosomes 7 and 9 was performed in workers occupationally exposed to low-dose benzene at a petroleum refinery. A micronucleus-centromere assay using a pan-centromeric probe was also performed to determine the origin of benzene-induced micronucleus (MN). Frequency of aneuploidy of chromosomes 7 and 9 was significantly higher among workers compared to the unexposed control group. Poisson regression analysis revealed that aneuploidy frequency of chromosome 7 or 9 was significantly associated with benzene level after adjusting for confounding variables such as age, smoking, alcohol intake, and duration of work (p = .042). Additionally, frequencies of MN and centromere-negative micronuclei (MNC-) were significantly higher in benzene-exposed workers compared to controls, while frequency of centromere-positive micronuclei (MNC+) was similar in both groups. In conclusion, aneuploidy of chromosomes 7 and 9 could be a useful biomarker to assess the effect of low-level benzene exposure, and benzene-induced MN originates from chromosome breaks rather than chromosome non-disjunction.