D-type cyclins and the DNA damage response

The cell cycle is strictly regulated to ensure the precise transmission of genetic information from one cell to its daughter cells. The DNA damage response is one of these regulatory pathways and is activated in response to both intrinsic and extrinsic genotoxic stress. Once activated, downstream signalling disrupts normal cell cycle progression, causing arrest or alternatively, cell death. One of the cell cycle proteins, cyclin D1, has been suggested to play a crucial role in integrating the cell cycle machinery and the DNA damage response. This investigation demonstrates that cyclin D1 expression responds in a dose-dependent manner to the UV-mimetic DNA damaging agent, 4-nitroquinolin 1-oxide (4NQO). Immunoblotting experiments revealed a biphasic response of cyclin D1 expression to 4NQO: cyclin D1 levels were largely unchanged following exposure of cells to low and high doses of 4NQO, whilst it was dramatically reduced after treatment with intermediate doses of 4NQO. The CDK inhibitor protein, p21, also responded in a dose-dependent manner, exhibiting elevated expression at low doses of 4NQO but was reduced or unchanged at intermediate and high doses, respectively. Exposure to IR or H2O2 triggered distinct responses by cyclin D1 and p21, indicating the likelihood that these DNA damaging agents activate distinct signalling pathways. Analyses of cellular responses by flow cytometry demonstrated that p21 expression primarily conferred resistance to damage-induced cell death whereas the level of cyclin D1 expression correlated with the mode of cell death. 4NQO-induced cyclin D1 downregulation was proteasome-dependent but was resistant to deregulation of the majority of the key damage-regulating proteins. This investigation also revealed that other members of D-type cyclin family, cyclins D2 and D3, also exhibit distinct responses to various DNA damaging agents. Preliminary observations indicated that responses of the three D-cyclins were not interdependent and they may play distinct roles in the DNA damage response

Gene-panel testing of breast and ovarian cancer patients identifies a recurrent RAD51C duplication

Gene-panel sequencing allows comprehensive analysis of multiple genes simultaneously and is now routinely used in clinical mutation testing of high-risk breast and ovarian cancer patients. However, only BRCA1 and BRCA2 are often analyzed also for large genomic changes. Here, we have analyzed 10 clinically relevant susceptibility genes in 95 breast or ovarian cancer patients with gene-panel sequencing including also copy number variants (CNV) analysis for genomic changes. We identified 12 different pathogenic BRCA1, BRCA2, TP53, PTEN, CHEK2, or RAD51C mutations in 18 of 95 patients (19%). BRCA1/2 mutations were observed in 8 patients (8.4%) and CHEK2 protein-truncating mutations in 7 patients (7.4%). In addition, we identified a novel duplication encompassing most of the RAD51C gene. We further genotyped the duplication in breast or ovarian cancer families (n=1149), in unselected breast (n=1729) and ovarian cancer cohorts (n=553), and in population controls (n=1273). Seven additional duplication carries were observed among cases but none among controls. The duplication associated with ovarian cancer risk (3/590 of all ovarian cancer patients, 0.5%, P=.032 compared with controls) and was found to represent a large fraction of all identified RAD51C mutations in the Finnish population. Our data emphasizes the importance of comprehensive mutation analysis including CNV detection in all the relevant genes.Peer reviewe

The mechanism of human basophil activation through the low affinity IgG receptor (Fc γ RⅡ) : Analysis of calcium mobilization using a new flow cytometric method

A new method for flow cytometric analysis of calcium mobilization in human peripheral blood basophils without prior purification was developed. The method is based on dual color analysis of centrifugation-enriched mononuclear cell populations using fluo-3 and phycoerythrin (PE)-conjugated CD2, CD14, CD16, CD19 monoclonal antibodies (mAb) to stain contaminated cells. This technique allows the detection of fluo-3 fluorescence as a measure of an increase in the cytoplasmic free calcium concentration ([Ca(2+)]i) while simulataneously discriminating PE-mAb-unlabelled basophils. To clarify whether the human peripheral blood basophil is activated through the low affinity IgG receptor, Fc γ RⅡ, as well as the high affinity IgE receptor, Fc ε　RⅠ, calcium mobilization after Fc γ RⅡ stimulation was analyzed by this method. After cross-linking of Fc γ RⅡ, transient [Ca(2+)]i elevation was observed but there was no apparent difference with interleukin-3(IL-3)-treated cells, and no significant histamine release was observed with or without short pre-incubation of IL-3. These findings suggest that the cross-linking of Fc γ RⅡ, not only Fc ε　RⅠ, can activate human basophils which may result in mediator release other than histamine

Adipocyte-specific protein tyrosine phosphatase 1B deletion increases lipogenesis, adipocyte cell size and is a minor regulator of glucose homeostasis

Peer reviewedPublisher PD

SETD2 loss-of-function promotes renal cancer branched evolution through replication stress and impaired DNA repair

The research leading to these results is supported by Cancer Research UK (XYG, RAB, EG, PM, PE, SG, C Santos, AJR, NM, PAB, AS and C Swanton), Breast Cancer Research Foundation (C Swanton and NK), Medical Research Council (ID: G0902275 to MG and C Santos; ID: G0701935/2 to AJR and C Swanton), the Danish Cancer Society (AMM, J Bartkova and J Bartek), the Lundbeck Foundation (R93-A8990 to J Bartek), the Ministry of the interior of the Czech Republic (grant VG20102014001 to MM and J Bartek), the National Program of Sustainability (grant LO1304 to MM and J Bartek), the Danish Council for Independent Research (grant DFF-1331-00262 to J Bartek), NIHR RMH/ICR Biomedical Research Centre for Cancer (JL), the EC Framework 7 (PREDICT 259303 to XYG, EG, PM, MG, TJ and C Swanton; DDResponse 259892 to J Bartek and J Bartkova and RESPONSIFY ID:259303 to C Swanton), UCL Overseas Research Scholarship (SG). C Swanton is also supported by the European Research Council, Rosetrees Trust and The Prostate Cancer Foundation. This research is supported by the National Institute for Health Research University College London Hospitals Biomedical Research Centre

The Hanshin-Awaji Great Earthquake in a Historical Perspective

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