Cell survival following radiation exposure requires miR-525-3p mediated suppression of ARRB1 and TXN1.

Background: microRNAs (miRNAs) are non-coding RNAs that alter the stability and translation efficiency of messenger RNAs. Ionizing radiation (IR) induces rapid and selective changes in miRNA expression. Depletion of the miRNA processing enzymes Dicer or Ago2 reduces the capacity of cells to survive radiation exposure.

Quantitative phosphoproteomic analysis of early alterations in protein phosphorylation by 2,3,7,8-tetrachlorodibenzo-p-dioxin.

A comprehensive quantitative analysis of changes in protein phosphorylation preceding or accompanying transcriptional activation by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in 5L rat hepatoma cells was performed using the SILAC approach. Following exposure of the cells to DMSO or 1 nM TCDD for 0.5 to 2 h, 5648 phosphorylated peptides corresponding to 2156 phosphoproteins were identified.

Evaluation of different biomarkers to predict individual radiosensitivity in an inter-laboratory comparison--lessons for future studies.

Radiotherapy is a powerful cure for several types of solid tumours, but its application is often limited because of severe side effects in individual patients. With the aim to find biomarkers capable of predicting normal tissue side reactions we analysed the radiation responses of cells from individual head and neck tumour and breast cancer patients of different clinical radiosensitivity in a multicentric study. Multiple parameters of cellular radiosensitivity were analysed in coded samples of peripheral blood lymphocytes (PBLs) and derived lymphoblastoid cell lines (LCLs) from 15 clinical radio-hypersensitive tumour patients and compared to age- and sex-matched non-radiosensitive patient controls and 15 lymphoblastoid cell lines from age- and sex- matched healthy controls of the KORA study.

A radiation-induced gene expression signature as a tool to predict acute radiotherapy-induced adverse side effects.

The majority of patients tolerate radiotherapy well, but some of them suffer from severe side effects. To find genes possibly predictive for radiosensitivity, mRNA profiles were generated before and 6h after in vitro irradiation with 5Gy. We analyzed lymphocytes from four head and neck and eight breast cancer patients with strong acute radiation toxicity and from 12 matching normal reacting patients in a blind study.

Ty1 integrase overexpression leads to integration of non-Ty1 DNA fragments into the genome of Saccharomyces cerevisiae.

The integrase of the Saccharomyces cerevisiae retrotransposon Ty1 integrates Ty1 cDNA into genomic DNA likely via a transesterification reaction. Little is known about the mechanisms ensuring that integrase does not integrate non-Ty DNA fragments. In an effort to elucidate the conditions under which Ty1 integrase accepts non-Ty DNA as substrate, PCR fragments encompassing a selectable marker gene were transformed into yeast strains overexpressing Ty1 integrase.

A novel function of Ubc13 in TNFR1 receptor activation.

The tumour necrosis factor receptor 1 (TNFR1) activates prosurvival pathways by induction of the NFkappaB pathway and induces cell death via apoptosis. The ubiquitin-conjugating enzyme, Ubc13, mediates the ubiquitylation-dependent formation of protein complexes crucial for the activation and regulation of both pathways. We describe a new role for Ubc13 in the regulation of TNFR1 activity after UV stimulation.

A novel function for the Mre11-Rad50-Xrs2 complex in base excision repair.

The Mre11/Rad50/Xrs2 (MRX) complex in Saccharomyces cerevisiae has well-characterized functions in DNA double-strand break processing, checkpoint activation, telomere length maintenance and meiosis. In this study, we demonstrate an involvement of the complex in the base excision repair (BER) pathway. We studied the repair of methyl-methanesulfonate-induced heat-labile sites in chromosomal DNA in vivo and the in vitro BER capacity for the repair of uracil- and 8-oxoG-containing oligonucleotides in MRX-deficient cells.

Dicentric chromosomes and gamma-H2AX foci formation in lymphocytes of human blood samples exposed to a CT scanner: a direct comparison of dose response relationships.

Experiments using the induction of dicentric chromosomes (dicentrics) as well as the gamma-H2AX foci formation in lymphocytes of blood samples from a healthy donor were performed to directly evaluate the radiation sensitivity of both biological endpoints. For computed tomography scans at dose levels from 0.025 to 1 Gy, a linear-quadratic dose-response relationship for dicentrics and a linear dose-response relationship for gamma-H2AX foci were obtained.

Multicentric investigation of ionising radiation-induced cell death as a predictive parameter of individual radiosensitivity.

In the present study, the predictive value of ionising radiation (IR)-induced cell death was tested in peripheral blood lymphocytes (PBLs) and their corresponding Epstein-Barr virus-transformed lymphoblastoid cell lines (LCLs) in an interlaboratory comparison. PBLs and their corresponding LCLs were derived from 15 tumour patients, that were considered clinically radiosensitive based on acute side-effects, and matched controls. Upon coding of the samples, radiosensitivity of the matched pairs was analysed in parallel in three different laboratories by assessing radiation-induced apoptotic and necrotic cell death using annexin V.

The DNA repair protein NBS1 influences the base excision repair pathway.

NBS1 fulfills important functions for the maintenance of genomic stability and cellular survival. Mutations in the NBS1 (Nijmegen Breakage Syndrome 1) gene are responsible for the Nijmegen breakage syndrome (NBS) in humans. The symptoms of this disease and the phenotypes of NBS1-defective cells, especially their enhanced radiosensitivity, can be explained by an impaired DNA double-strand break-induced signaling and a disturbed repair of these DNA lesions.

Reduced expression of SRC family kinases decreases PI3K activity in NBS1-/- lymphoblasts.

SRC family kinases (SFKs) are involved in the activation of phosphatidylinositol-3-kinase (PI3K). In addition, the activity of this lipid kinase can be regulated by the DNA repair protein NBS1. Here, we describe a disturbed expression of some members of the non-receptor tyrosine kinase family in lymphoblastoid cell lines generated from cells of Nijmegen breakage syndrome (NBS) patients.

NBS1 is required for IGF-1 induced cellular proliferation through the Ras/Raf/MEK/ERK cascade.

NBS1 is a member of the Mre11-Rad50-NBS1 complex, which plays a role in cellular responses to DNA damage and the maintenance of genomic stability. Transgenic mice models and clinical symptoms of NBS patients have shown that NBS1 exerts pleiotropic actions on the growth and development of mammals. The present study showed that after repression of endogenous NBS1 levels using short interfering RNA, hTERT-RPE cells demonstrated impaired proliferation and a poor response to IGF-1.

Non-conservative homologous recombination in human B lymphocytes is promoted by activation-induced cytidine deaminase and transcription.

During secondary immunoglobulin (Ig) diversification in vertebrates, the sequence of the variable region of Ig genes may be altered by templated or non-templated mechanisms. In both cases, cytidine deamination by activation-induced cytidine deaminase (AID) in the transcribed Ig loci leads to DNA lesions, which are repaired by conservative homologous recombination (HR) during Ig gene conversion, or by non-templated mutagenesis during somatic hypermutation. The molecular basis for the differential use of these two pathways in different species is unclear.

Regulation of double-stranded DNA gap repair by the RAD6 pathway.

The RAD6 pathway allows replication across DNA lesions by either an error-prone or error-free mode. Error-prone replication involves translesion polymerases and requires monoubiquitylation at lysine (K) 164 of PCNA by the Rad6 and Rad18 enzymes. By contrast, the error-free bypass is triggered by modification of PCNA by K63-linked polyubiquitin chains, a reaction that requires in addition to Rad6 and Rad18 the enzymes Rad5 and Ubc13-Mms2.

Xrs2 facilitates crossovers during DNA double-strand gap repair in yeast.

Xrs2 is a member of the MRX complex (Mre11/Rad50/Xrs2) in Saccharomyces cerevisiae. In this study we demonstrate the important role of the MRX complex and in more detail of Xrs2 for the repair of radiation-induced chromosomal double-strand breaks by pulsed field gel electrophoresis. By using a newly designed in vivo plasmid-chromosome recombination system, we could show that gap repair efficiency and the association with crossovers were reduced in the MRX null mutants, but repair accuracy was unaffected.

Enhanced CD95-mediated apoptosis contributes to radiation hypersensitivity of NBS lymphoblasts.

The molecular causes for enhanced radiosensitivity of Nijmegen Breakage Syndrome cells are unclear, especially as repair of DNA damage is hardly impeded in these cells. We clearly demonstrate that radiation hypersensitivity is accompanied by enhanced gamma-radiation-induced apoptosis in NBS1 deficient lymphoblastoid cell lines. Differences in the apoptotic behavior of NBS1 (-/-) and NBS1 (+/-) cells are not due to an altered p53 stabilization or phosphorylation in NBS1 (-/-) cells.

The rate of extrachromosomal homologous recombination within a novel reporter plasmid is elevated in cells lacking functional ATM protein.

Homologous recombination between identical stretches of DNA depends on the coordinated action of many tightly regulated proteins. Cellular defects in homologous recombination are strongly associated with increased genomic instability and tumorigenesis. In cells of the cancer-prone syndrome ataxia telangiectasia (A-T), increased intrachromosomal recombination has been demonstrated, while extrachromosomal recombination has been discussed controversially.

Spontaneous homologous recombination is decreased in Rad51C-deficient hamster cells.

The Chinese hamster cell mutant, CL-V4B that is mutated in the Rad51 paralog gene, Rad51C (RAD51L2), has been described to exhibit increased sensitivity to DNA cross-linking agents, genomic instability, and an impaired Rad51 foci formation in response to DNA damage. To directly examine an effect of the Rad51C protein on homologous recombination (HR) in mammalian cells, we compared the frequencies and rates of spontaneous HR in CL-V4B cells and in parental wildtype V79B cells, using a recombination reporter plasmid in host cell reactivation assays. Our results demonstrate that HR is reduced but not abolished in the CL-V4B mutant.

Tumor-associated E-cadherin mutations do not induce Wnt target gene expression, but affect E-cadherin repressors.

E-cadherin is a cell-cell adhesion molecule and tumor invasion suppressor gene that is frequently altered in human cancers. It interacts through its cytoplasmic domain with beta-catenin which in turn interacts with the Wnt (wingless) signaling pathway. We have compared the effects of different tumor-derived E-cadherin variants with those of normal E-cadherin on Wnt signaling and on genes involved in epithelial mesenchymal transition.

Promoter-trapping in Saccharomyces cerevisiae by radiation-assisted fragment insertion.

Non-homologous insertion (NHI) of DNA fragments into genomic DNA is a method widely used in insertional mutagenesis screens. In the yeast Saccharomyces cerevisiae, the efficiency of NHI is very low. Here we report that its efficiency can be increased by gamma-irradiation of recipient cells at the time of transformation.