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The meiosis-specific synaptonemal complex protein SYCP3 has been reported to be aberrantly expressed in tumours. However, in contrast to its well-defined function in meiosis, its possible role in mitotic cells is entirely unknown. Here, we show that SYCP3 is expressed in a range of primary tumours and that it impairs chromosomal integrity in mitotic cells. Expression of SYCP3 inhibits the homologous recombination (HR) pathway mediated by RAD51, inducing hypersensitivity to DNA-damaging agents such as a poly(ADP-ribose) polymerase (PARP) inhibitor and chromosomal instability. SYCP3 forms a complex with BRCA2 and inhibits its role in HR. These findings highlight a new mechanism for chromosomal instability in cancer and extend the range of PARP-inhibitor sensitive tumours to those expressing SYCP3. [ABSTRACT FROM AUTHOR]

Chromosome translocations induced by DNA damaging agents, such as ionizing radiation and certain chemotherapies, alter genetic information resulting in malignant transformation. Abrogation or loss of the ataxia-telangiectasia mutated (ATM) protein, a DNA damage signaling regulator, increases the incidence of chromosome translocations. However, how ATM protects cells from chromosome translocations is still unclear. Chromosome translocations involving the MLL gene on 11q23 are the most frequent chromosome abnormalities in secondary leukemias associated with chemotherapy employing etoposide, a topoisomerase II poison. Here we show that ATM deficiency results in the excessive binding of the DNA recombination protein RAD51 at the translocation breakpoint hotspot of 11q23 chromosome translocation after etoposide exposure. Binding of Replication protein A (RPA) and the chromatin remodeler INO80, which facilitate RAD51 loading on damaged DNA, to the hotspot were also increased by ATM deficiency. Thus, in addition to activating DNA damage signaling, ATM may avert chromosome translocations by preventing excessive loading of recombinational repair proteins onto translocation breakpoint hotspots. [ABSTRACT FROM AUTHOR]

In human somatic cells, homologous recombination is a rare event. To facilitate the targeted modification of the genome for research and gene therapy applications, efforts should be directed toward understanding the molecular mechanisms of homologous recombination in human cells. Although human genes homologous to members of the RAD52 epistasis group in yeast have been identified, no genes have been demonstrated to play a role in homologous recombination in human cells. Here, we report that RAD54B plays a critical role in targeted integration in human cells. Inactivation of RAD54B in a colon cancer cell line resulted in severe reduction of targeted integration frequency. Sensitivity to DNA-damaging agents and sister-chromatid exchange were not affected in RAD54B-deficient cells. Parts of these phenotypes were similar to those of Saccharomyces cerevisiae tid1/rdh54 mutants, suggesting that RAD54B may be a human homolog of TID1/RDH54. In yeast, TID1/RDH54 acts in the recombinational repair pathway via roles partially overlapping those of RAD54. Our findings provide the first genetic evidence that the mitotic recombination pathway is functionally conserved from yeast to humans. [ABSTRACT FROM AUTHOR]

Histone modifications are catalyzed and recognized by specific proteins to regulate dynamic DNA metabolism processes. NSD2 is a histone H3 lysine 36 (H3K36)‐specific methyltransferase that is associated with both various transcription regulators and DNA repair factors. Specifically, it has been implicated in the repair of DNA double‐strand breaks (DSBs); however, the role of NSD2 during DSB repair remains enigmatic. Here, we show that NSD2 does not accumulate at DSB sites and that it is not further mobilized by DSB formation. Using three different DSB repair reporter systems, which contained the endonuclease site in the active thymidine kinase gene (TK) locus, we demonstrated separate dose‐dependent effects of NSD2 on homologous recombination (HR), canonical‐non‐homologous end joining (c‐NHEJ), and non‐canonical‐NHEJ (non‐c‐NHEJ). Endogenous NSD2 has a role in repressing non‐c‐NHEJ, without affecting DSB repair efficiency by HR or total NHEJ. Furthermore, overexpression of NSD2 promotes c‐NHEJ repair and suppresses HR repair. Therefore, we propose that NSD2 has functions in chromatin integrity at the active regions during DSB repair. [ABSTRACT FROM AUTHOR]

Acute promyelocytic leukemia (APL) has been characterized by 15;17 chromosomal translocation, which involves the retinoic acid receptor alpha (RARA) gene on chromosome 17 and the PML gene on chromosome 15, An extremely restricted region (ERR) of 50bps within the second intron of the RARA gene was identified as the cluster region of breakpoints by sequencing analyses. ERR was tested by in vitro transfection-recombination assay, and was shown to be the recombination hot spot. In this study, presence of DNA binding proteins to the 148bps DNA fragment which contains ERR was confirmed by gel-mobility shift analysis in the nuclear extract of NIH3T3 cells and human leukemia cell lines. Furthermore, in vitro study with the mouse sarcoma cell lines using the recombination reporter plasmid containing ERR showed that ERR might be involved in the homologous recombination in addition to the illegitimate recombination. The DNA binding proteins specific to ERR might play an important role in chromosome translocation. [ABSTRACT FROM AUTHOR]

According to an analysis of the Life Span Study cohort data conducted by the Radiation Effects Research Foundation in Hiroshima and Nagasaki, the sex-averaged excess relative risk (ERR) of all solid cancers was 0.42 Gy-Eq-1. On the other hand, analysis of the atomic bomb survivors (ABS) cohort data at Hiroshima University indicated the ERR value was 0.28 Gy-Eq-1 in Hiroshima. In both cases, initial radiation doses were derived from the dosimetry system DS02, in which the relative biological effectiveness (RBE) of neutrons was assumed to be a constant value of 10. To clarify the validity of the RBE, the authors investigated the possibility of different contributions of neutrons by using the ABS. Although there were no statistically significant differences among the estimated value of RBE (=65) and the ordinal value (=10), the corresponding ERR decreased by 30%, which might affect the interpretation of radiation health assessments.

Genetic information encoded in chromosomal DNA is challenged by intrinsic and exogenous sources of DNA damage. DNA double-strand breaks (DSBs) are extremely dangerous DNA lesions. RAD51 plays a central role in homologous DSB repair, by facilitating the recombination of damaged DNA with intact DNA in eukaryotes. RAD51 accumulates at sites containing DNA damage to form nuclear foci. However, the mechanism of RAD51 accumulation at sites of DNA damage is still unclear. Post-translational modifications of proteins, such as phosphorylation, acetylation and ubiquitylation play a role in the regulation of protein localization and dynamics. Recently, the covalent binding of small ubiquitin-like modifier (SUMO) proteins to target proteins, termed SUMOylation, at sites containing DNA damage has been shown to play a role in the regulation of the DNA-damage response. Here, we show that the SUMOylation E2 ligase UBC9, and E3 ligases PIAS1 and PIAS4, are required for RAD51 accretion at sites containing DNA damage in human cells. Moreover, we identified a SUMO-interacting motif (SIM) in RAD51, which is necessary for accumulation of RAD51 at sites of DNA damage. These findings suggest that the SUMO-SIM system plays an important role in DNA repair, through the regulation of RAD51 dynamics.

Ionizing radiation has been shown to induce various types of chromosomal DNA damages. Among these DNA damages, DNA double strand breaks(DSBs) are the most severe damages resulting in cell death or chromosome abnormalities. Proteins associated with DNA repair, such as phosphorylated form of histone H2AX, a histone variant of H2A, and a DNA recombinase RAD51, has been shown to form radiation-induced repair foci at sites containing DNA damage. Reorganization of damaged chromatin by protein modifications or exchange of histones has been shown to play an important role in the formation of radiation induced repair foci.

Objective: Estrogen regulates the expression of epithelial Na(+) channel (ENaC) and cystic fibrosis transmembrane conductance regulator (CFTR). Our purpose was to assess the effects of raloxifene analog LY117018 on the expression of ENaC and CFTR in ovariectomized mice., Study Design: Three groups of 5 female ovariectomized mice were treated with 17beta-estradiol benzoate (E2), LY117018 (LY), or vehicle, respectively, for 4-12 weeks. Effects on the messenger ribonucleic acid expression levels of ENaC and CFTR channels in the uterus were studied using real-time reverse transcriptase-polymerase chain reaction., Results: E2 treatment induced CFTR expression, repressed ENaC expression and resulted in fluid accumulation in the uterus. In contrast, LY induced CFTR expression, did not repress ENaC expression, and caused no fluid accumulation., Conclusion: Estradiol and LY117018 differentially regulate the expression of CFTR and ENaC in ovariectomized mouse uterus. This finding suggests that uterine fluid accumulation can be controlled mainly by targeting the ENaC.

BACH2 is a B-cell-specific transcription repressor and is also know as a tumor suppressor in B cell malignancy. Expression of BACH2 is induced in BCR-ABL positive lymphoid cell lines including BV173 by imatinib, a molecular targeting agent for the treatment of chronic myeloid leukemia (CML). Here we show that the activity of the BACH2 gene is related to the nuclear positioning of the gene loci. We examined the spatial association of the BACH2 gene with the centromeric heterochromatin, a transcriptionally repressive subnuclear compartment, by comparing cells with low (BV173 and K562) and high (NAMALWA) levels of BACH2 mRNA. The BACH2 gene was located closer to the centromeric heterochromatin in BV173 and K562 cells as compared to NAMALWA cells. In BV173 cells, the BACH2-centromere distance increased after imatinib treatment to levels similar to those in NAMALWA cells. We also found that diethylmaleate, an oxidative stressor, enhanced the antiproliferative effect of imatinib in only BV173 cells. Since BACH2 induces apoptosis by oxidative stress, these observations suggest that down-regulation of the BACH2 gene through the interaction with centromeric heterochromatin would take part in leukomogenesis of BCR-ABL positive lymphoid leukemia., (Copyright 2006 Wiley-Liss, Inc.)

Small Maf proteins serve as dual-function transcription factors through an exchange of their heterodimerization partners. For example, as heterodimers with hematopoietic cell-specific p45 NF-E2 or NF-E2-related factors (Nrf), they activate the beta-globin or antioxidative stress enzyme heme oxygenase 1 (HO-1) genes, respectively. In contrast, together with Bach1, they repress these same genes. However, the signals leading to this partner exchange are not known. Using chromatin immunoprecipitation assays in NIH 3T3 cells, we show that heme, an inducer of ho-1, promotes displacement of Bach1 from the MafK-occupied ho-1 enhancers, which is followed by Nrf2 binding to these elements. Whereas histone H3 at the ho-1 enhancers and promoter is hyperacetylated irrespective of gene activity, exposure of cells to heme results in de novo hyperacetylation and hypermethylation of histone H3 in the transcribed region. These data indicate that, under normal conditions, the chromatin structure of ho-1 is in a preactivation state, but transcription is repressed by Bach1. Heme induces switching of Maf dimers, resulting in ho-1 expression. Heme also promotes displacement of Bach1 from the beta-globin locus control region without affecting MafK binding in murine erythroleukemia cells. Thus, heme functions as a signaling molecule for gene expression in higher eukaryotes.

Whether chromosomes maintain their nuclear positions during interphase and from one cell cycle to the next has been controversially discussed. To address this question, we performed long-term live-cell studies using a HeLa cell line with GFP-tagged chromatin. Positional changes of the intensity gravity centers of fluorescently labeled chromosome territories (CTs) on the order of several microm were observed in early G1, suggesting a role of CT mobility in establishing interphase nuclear architecture. Thereafter, the positions were highly constrained within a range of approximately 1 microm until the end of G2. To analyze possible changes of chromosome arrangements from one cell cycle to the next, nuclei were photobleached in G2 maintaining a contiguous zone of unbleached chromatin at one nuclear pole. This zone was stably preserved until the onset of prophase, whereas the contiguity of unbleached chromosome segments was lost to a variable extent, when the metaphase plate was formed. Accordingly, chromatin patterns observed in daughter nuclei differed significantly from the mother cell nucleus. We conclude that CT arrangements were stably maintained from mid G1 to late G2/early prophase, whereas major changes of CT neighborhoods occurred from one cell cycle to the next. The variability of CT neighborhoods during clonal growth was further confirmed by chromosome painting experiments.