Search results for "REPLICATION"
showing 10 items of 489 documents
Single molecule localization microscopy of the distribution of chromatin using Hoechst and DAPI fluorescent probes.
2014
Several approaches have been described to fluorescently label and image DNA and chromatin in situ on the single-molecule level. These superresolution microscopy techniques are based on detecting optically isolated, fluorescently tagged anti-histone antibodies, fluorescently labeled DNA precursor analogs, or fluorescent dyes bound to DNA. Presently they suffer from various drawbacks such as low labeling efficiency or interference with DNA structure. In this report, we demonstrate that DNA minor groove binding dyes, such as Hoechst 33258, Hoechst 33342, and DAPI, can be effectively employed in single molecule localization microscopy (SMLM) with high optical and structural resolution. Upon ill…
Origin of the integrin-mediated signal transduction. Functional studies with cell cultures from the sponge Suberites domuncula
1999
Sponges (phylum Porifera) represent the phylogenetically oldest metazoan animals. Recently, from the marine sponge Geodia cydonium a first cDNA encoding a putative integrin receptor molecule was isolated. In the present study basic functional experiments have been conducted to test the hypothesis that in sponges integrin polypeptides also function as adhesion molecules and as outside-in signaling molecules. The sponge Suberites domuncula has been used for the experiments because from this sponge only has a cell culture been established. Here we report that aggregation factor (AF)-mediated cell-cell adhesion is blocked by the RGDS peptide which is known to interact with beta integrin. Both R…
TWJ-Screen: an isothermal screening assay to assess ligand/DNA junction interactions in vitro
2017
International audience; The quest for chemicals able to operate at selected genomic loci in a spatiotemporally controlled manner is desirable to create manageable DNA damages. Mounting evidence now shows that alternative DNA structures, including G-quadruplexes and branched DNA (or DNA junctions), might hamper proper progression of replication fork, thus triggering DNA damages and genomic instability. Therefore, small molecules that stabilize these DNA structures are currently scrutinized as a promising way to create genomic defects that cannot be dealt with properly by cancer cells. While much emphasis has been recently given to G-quadruplexes and related ligands, we report herein on three…
Measurement of Lymphocyte Proliferation: Critical Analysis of Radioactive and Photometric Methods
1992
Different methods of lymphocyte proliferation are compared to identify a non-radioactive alternative to 3H-thymidine-test. The enzymatic assays evaluating the turnover of mitochondrial dehydrogenases (MTT-test) and lysosomal hexosaminidase (NAG-test) proved not sensitive enough to substitute for 3H-thymidine incorporation. The incorporation of the nucleotide analog 5-bromodeoxyuridine (BrdU) can be exploited using an ELISA-system (enzyme linked immunosorbent assay) employing a monoclonal anti-BrdU antibody to measure cell proliferation. An optimized test protocol of the BrdU-ELISA which fulfills the requirements for a sensitive and practicable non-radioactive alternative to 3H-thymidine-tes…
How DNA lesions are turned into powerful killing structures: Insights from UV-induced apoptosis
2008
Mammalian cells treated with ultraviolet (UV) light provide one of the best-known experimental systems for depicting the biological consequences of DNA damage. UV irradiation induces the formation of DNA photoproducts, mainly cyclobutane pyrimidine dimers (CPDs) and (6-4) pyrimidine-pyrimidone photoproducts [(6-4)PPs], that drastically impairs DNA metabolism, culminating in the induction of cell death by apoptosis. While CPDs are the most important apoptosis-inducing lesions in DNA repair proficient cells, recent data indicates that (6-4)PPs also signals for apoptosis in DNA repair deficient cells. The toxic effects of these unrepaired DNA lesions are commonly associated with transcription …
Microenvironmental adaptation of experimental tumours to chronic vs acute hypoxia
2004
This study investigated long-term microenvironmental responses (oxygenation, perfusion, metabolic status, proliferation, vascular endothelial growth factor (VEGF) expression and vascularisation) to chronic hypoxia in experimental tumours. Experiments were performed using s.c.-implanted DS-sarcomas in rats. In order to induce more pronounced tumour hypoxia, one group of animals was housed in a hypoxic atmosphere (8% O(2)) for the whole period of tumour growth (chronic hypoxia). A second group was acutely exposed to inspiratory hypoxia for only 20 min prior to the measurements (acute hypoxia), whereas animals housed under normal atmospheric conditions served as controls. Acute hypoxia reduced…
The APC/C cofactor Cdh1 prevents replicative stress and p53-dependent cell death in neural progenitors
2013
The E3-ubiquitin ligase APC/C-Cdh1 is essential for endoreduplication but its relevance in the mammalian mitotic cell cycle is still unclear. Here we show that genetic ablation of Cdh1 in the developing nervous system results in hypoplastic brain and hydrocephalus. These defects correlate with enhanced levels of Cdh1 substrates and increased entry into the S phase in neural progenitors. However, cell division is prevented in the absence of Cdh1 due to hyperactivation of cyclin-dependent kinases, replicative stress, induction of p53, G2 arrest and apoptotic death of these progenitor cells. Concomitant ablation of p53 rescues apoptosis but not replicative stress, resulting in the presence of …
Are the leukocyte telomere length attrition and telomerase activity alteration potential predictor biomarkers for sporadic TAA in aged individuals?
2014
A large variability in occurrence, complications, and age/gender manifestations characterizes individual susceptibility of sporadic thoracic aortic aneurysms (TAA), even in subjects with the same risk factor profiles. The reasons are poorly understood. On the other hand, TAA pathophysiology mechanisms remain unclear than those involved in abdominal aorta aneurysms. However, recent evidence is suggesting a crucial role of biological ageing in inter-individual risk variation of cardiovascular diseases, including sporadic TAA. Biological age rather than chronological age is a better predictor of vascular risk. Relevant assumptions support this concept. In confirming this evidence and our preli…
Asynchronous replication dynamics of imprinted and non-imprinted chromosome regions in early mouse embryos.
2008
We have used interphase FISH to analyze the replication behavior of four imprinted chromosome regions (Snrpn, Zim1-Peg3, Dlk1-Gtl2, and Igf2r) and five non-imprinted regions in mouse one-cell to morula-stage embryos and embryonic fibroblasts. In general, imprinted chromosome regions showed the expected asynchronous pattern of replication throughout all analyzed stages of preimplantation development and in differentiated cells. The Dlk1-Gtl2 locus which is not expressed and Igf2r which is biallelically expressed in early embryos showed a relaxation of replication asynchrony at the morula stage. Asynchronous replication in zygotes and two-cell embryos was not specific to imprinted regions. Th…
Chromosomal instability, reproductive cell death and apoptosis induced by O6-methylguanine in Mex−, Mex+ and methylation-tolerant mismatch repair com…
1998
O6-Methylguanine (O6-MeG) is induced in DNA by methylating environmental carcinogens and various cytostatic drugs. It is repaired by O6-methylguanine-DNA methyltransferase (MGMT). If not repaired prior to replication, the lesion generates gene mutations and leads to cell death, sister chromatid exchanges (SCEs), chromosomal aberrations and malignant transformation. To address the question of how O6-MeG is transformed into genotoxic effects, isogenic Chinese hamster cell lines either not expressing MGMT (phenotypically Mex-), expressing MGMT (Mex+) or exhibiting the tolerance phenotype (Mex-, methylation resistant) were compared as to their clastogenic response. Mex- cells were more sensitiv…