3,243 research outputs found
Aurora B inhibitors promote RB hypophosphorylation and senescence independent of p53-dependent CDK2/4 inhibition
No full textAbstract Aurora B kinase (AURKB) inhibitors have been trialled in a range of different tumour types but are not approved for any indication. Expression of the human papilloma virus (HPV) oncogenes and loss of retinoblastoma (RB) protein function has been reported to increase sensitivity to AURKB inhibitors but the mechanism of their contribution to sensitivity is poorly understood. Two commonly reported outcomes of AURKB inhibition are polyploidy and senescence, although their relationship is unclear. Here we have investigated the major cellular targets of the HPV E6 and E7, p53 and RB, to determine their contribution to AURKB inhibitor induced polyploidy and senescence. We demonstrate that polyploidy is a universal feature of AURKB inhibitor treatment in all cell types including normal primary cells, but the subsequent outcomes are controlled by RB and p53. We demonstrate that p53 by regulating p21 expression is required for an initial cell cycle arrest by inhibiting both CDK2 and CDK4 activity, but this arrest is only triggered after cells have undergone two failed mitosis and cytokinesis. However, cells can enter senescence in the absence of p53. RB is essential for AURKB inhibitor-induced senescence. AURKB inhibitor induces rapid hypophosphorylation of RB independent of inhibition of CDK2 or CDK4 kinases and p53. This work demonstrates that p53 activation determines the timing of senescence onset, but RB is indispensable for senescence
Liquid structure of Rb-Hg alloys studied by neutron diffraction
No full textThe structures of liquid Rb–Hg alloys were studied as a function of composition by neutron diffraction. In the intermediate Rb concentration range, the obtained structure factors show a small prepeak, which may be an evidence of the formation of Hg polyanion units in liquids. The Reverse Monte Carlo (RMC) analysis was applied to separate the total radial distribution function into the corresponding partial radial distribution functions. Up to 10 at.% Rb, no obvious changes are found for the first peak position of the partial radial distribution functions of the Hg–Hg pair and that of the Hg–Rb pair. The first peak position between the Hg–Rb pairs increases above 20 at.% Rb. In addition to the first peak, a subpeak between Hg–Hg pairs can be seen in the large distance. At 60 at.% Rb, the nearest neighbor distance between Hg atoms shows the closest value in the concentration range studied. These results indicate that with the progress of charge transfer the solvation structure in the dilute Rb concentration range changes into the structure containing polyanions composed of Hg species
Fractional Edge Cover Number of Model RB
No full textModel RB is a random constraint satisfaction problem with a growing domain size, which exhibits exact phase transition phenomena. Many hard instances with planted solutions can be generated via Model RB, to be used as benchmarks for algorithmic competitions and researches. In the past, some structural parameters of constraint hypergraphs are analyzed to show hardness of Model RB, such as hinge width, decycling number, treewidth, and hypertree width. In this paper, one more structural parameter of constraint hypergraphs of Model RB, namely the fractional edge cover number, is analyzed. We show upper and lower bounds on the fractional edge cover number of Model RB. In particular, the fractional edge cover number of Model RB is shown to be asymptotically linear in the number of variables, like hinge width, decycling number, treewidth and hypertree width. These results together provide further evidences on the hardness of Model RB.EICPCI-S(ISTP)[email protected]
Developing an Immunofluorescence Assay for Detecting Rb and phospho-Rb on Circulating Tumor Cells in Breast Cancer
No full textEndocrine therapies (ET) such as tamoxifen, fulvestrant, and aromatase inhibitors (AIs) are the standard-of-care first-line treatment in majority of estrogen receptor (ER)-positive breast cancers (BC). Recent clinical studies using cyclin-dependent kinase 4 and 6 inhibitors (CDK4/6i) plus AIs or fulvestrant have shown significant improvement in survival outcomes in patients with ER+ metastatic BC compared to standalone ETs. CDK4/6i exert their action by inhibiting the phosphorylation of retinoblastoma (Rb) protein and consequently inducing cell cycle arrest. However, not all patients respond to this combination therapy and those, who initially respond, eventually develop resistance. Emerging studies suggest that the intrinsic resistance to CDK4/6i could be due to the loss of Rb or its mutations. Therefore, CDK4/6i resistance can be evaluated by measuring expression of total and phospho-Rb and Rb mutations. However, repeated biopsies to evaluate biomarkers in tumors is not feasible in patients. In this research, we aimed to develop an immunofluorescence assay to evaluate the expression of Rb and phospho-Rb using circulating tumor cells (CTCs) which can help predict response and resistance to CDK4/6i. CTCs serve as representative of the tumor bulk in patients and animal models and allow for less-invasive, frequent blood collection and real-time monitoring of treatment response. MCF7 cells treated with vehicle or abemaciclib (500 nM) for 48 hours were spiked into blood from non-tumor bearing mice. The MCF7 cells-spiked blood was processed using ScreenCell® device and the cells were transferred to slides and stained with DAPI, pan-cytokeratin, CD45, estrogen receptor, Rb and phospho-Rb. Tumor cells were defined as pan-cytokeratin-positive, CD45-negative, and nuclear stain-positive cells. Various antibody combinations were examined to increase the sensitivity of the IF assay for individual markers as well as the multiplexed assay. We also developed quantitative assessment approach to detect per-cell intensity of various markers. Treatment with abemaciclib reduced the intensity of phospho/Total Rb from 2.6 +/- 0.6 to 0.8 +/- 0.2 units (p < 0.05, t-test, n=8-9) in the vehicle-treated samples. There was no significant difference in the Rb intensity between the treatment groups. Ongoing studies focus on validation of the assay using preclinical models and clinical samples.Pharmacy Practice and Translational Research, Department ofHonors Colleg
Inhibition of Aurora B kinase activity triggers senescence that can be bypassed by blocking p53 and RB function, promoting replication stress
No full textAurora B kinase has a major role in regulating progression through mitosis and partitioning the replicated genome at exit from mitosis and cytokinesis. We have previously reported that Aurora A and B inhibitors selectively targeted HPV-driven tumours, and that the HPV E7 oncogene is a major determinant of this selectivity. Another group has shown that sensitivity to Aurora B inhibitor AZD2811 is dependent on loss of the retinoblastoma protein (RB). Here we have investigated the outcomes of inhibition of Aurora B using selective and pan Aurora inhibitors. We show that inhibition of Aurora A and B promotes more cell death, although a subset of cells are protected from the cytotoxic effects as a consequence of Aurora A inhibition slowing the cell cycle. Aurora B inhibition promotes senescence, although this requires at least two cycles of failed cytokinesis in all cell types, including primary fibroblasts. Loss of RB function either by HPV E7 or CDK4 R24C mutant over-expression, and loss of p53 function have somewhat different effects on the outcomes of Aurora B inhibition, although both reduced the proportion of cells that entered senescence. Loss of either also promoted replication stress which was not observed in RB and p53 proficient cells, the level of replication stress was not enhanced by Aurora A co-inhibition. Selective Aurora B inhibitor was also less toxic to proliferating PBMC derive CD3+ T cells than the pan Aurora inhibitor. Together these data indicate that short term treatment with Aurora B selective inhibitor is sufficient to promote cell cycle arrest and senescence in RB and p53 proficient cells, although proliferating T cells appear to tolerate this. RB and p53 pathway defective cells are less sensitive to this senescence trigger, but undergo replication stress.No Full Tex
MILLIMETER-WAVE SPECTROSCOPY OF COLD RB ATOMS
No full textAuthor Institution: Department of Physics, University of Virginia, McCormick Road, Charlottesville, Virginia 22903Cold Rb atoms were prepared by magneto-optical trap. Millimeter-wave has been used to drive nd to (n-2)f one-photon and nd to (n-1)g () two-photon transitions. Quantum defects of f and g states of Rb were calculated. Full analyses will be presented. }
Interpretation and the Problem of the Intention of the Author, by Burhanetir Tatar
No full textBurhanetir Tatar, Interpretation and the Problem of the Intention of the Author: H.G. Gadamer vs E.D. Hirsh, The Council for Research in Values and Philosophy, 199
Rb interactome data and its modulations during cell cycle progression in HEK 293 cells
No full textThe Rb protein is a tumor suppressor protein that regulates the key G1S checkpoint consequently blocking the progression of cell cycle into S-phase. Despite its pertinent role in cell cycle regulation, comprehensive information on its interacting partners across cell cycle progression is lacking. Here, we aim to submit a comprehensive set of Rb interactors as the cell progresses from G0 through G1 and S into G2 phase in HEK 293 cell line. Affinity purification of HA-tagged Rb protein along with its interactors was analyzed by mass spectrometry (AP-MS). SILAC labeling enabled differentiation of Rb interactors in different cell cycle stages as well as their quantification - G0 cells were labeled with light labels of lysine and arginine (K0R0), cells in G1S transition were labeled with heavy labels (K8R10) while the G2 cells were labeled with medium labels (K6R6). LC-MS/MS analysis resulted in 6 wiff files which were submitted to protein pilot software for peptide identification and quantification. Here we submit the dataset which clearly captures the changing interacting partners of the Rb protein as the cell cycle progressed from G0 through G1S checkpoint into G2 phase. Data is publicly available via ProteomeXchange with identifier PXD007708. Keywords: Rb, Cell cycle, Interactome, SILAC, AP-MS, HEK 293 cell
Aneuploidy in spermatids of Robertsonian (Rb) chromosome heterozygous mice
No full text© 2014, The Author(s). Rb translocations are chromosomal rearrangements frequently found in natural populations of the house mouse Mus musculus domesticus. The standard diploid karyotype of the house mouse consisting of 40 telocentric chromosomes may be reduced by the emergence of metacentric Rb chromosomes. Multiple simple Rb heterozygotes form trivalents exhibiting higher anaphase nondisjunction frequency and consequently higher number of unbalanced gametes than in normal males. This work will attempt to establish whether frequencies of aneuploidy observed in heterozygote spermatids of the house mouse M. musculus domesticus show differences in chromosomes derived from different trivalents. Towards this goal, the number and distribution frequency of aneuploidy was assessed via FISH staining of specific chromosomes of spermatids derived from 2n = 32 individuals. Our results showed that for a given set of target chromosomes, 90 % of the gametes were balanced, resulting from alternate s
Synthesis and Characterization of Multiple-Cation Rb(MAFA)PbI3 Perovskite Single Crystals
No full textWe synthesized multiple-cation Rb(MAFA)PbI3 perovskite single crystals for the first time. The effect of Rb+ substitution was systemically investigated, and the addition of 1.5 M 5% RbI was the optimum condition to obtain high-quality Rb(MAFA)PbI3 single crystals. Lattice shrinkage occurred in the Rb(MAFA)PbI3 single crystal because of the small ionic radius of Rb+, resulting in blue-shifted absorption and photoluminescence (PL) peaks. The 1.5 M 5% RbI-added (MAFA)PbI3 single crystal showed the longest carrier lifetime of 18.35 ns, exhibiting the highest photoresponse than other crystals. We believe that this work will provide a basic insight into the mixed-cation perovskite single crystals for the future optoelectronic applications. © The Author(s) 201
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