ABSTRACT
Following prolonged mitotic spindle disruption by microtubule poisons, mammalian cells delay their entry into anaphase, then progressively slip out of mitosis and become tetraploid. Normal cells then stop cycling before S-phase onset, but the mechanisms underlying this arrest are still unclear. Here we show that a double block prevents endo-reduplication. First, cells that exit mitosis without a functional microtubule network are driven toward G0. Reconstitution of the network unmasks a second block that relies on DNA double-strand breaks occurring early in the G1 phase that follows the mitotic block. We propose that a stress signal elicited upon mitotic impairment triggers breakage, which couples the leaky spindle checkpoint to the stringent DNA damage response. Consistent with this finding, cells defective for the damage response continue cycling and acquire, within a single cell cycle, both chromosome rearrangements and abnormal chromosome numbers that remarkably mimic the complex genetic hallmark of tumorigenesis.
Subject(s)
Chromosomes , DNA Damage , Mitosis/physiology , Ploidies , Adenocarcinoma/pathology , Anaphase , Animals , Antineoplastic Agents/pharmacology , Cells, Cultured , Colonic Neoplasms/pathology , Cricetinae , Cricetulus , DNA Breaks, Double-Stranded , Fibroblasts/physiology , G1 Phase/genetics , Gene Rearrangement , Humans , Immunoblotting , In Situ Hybridization, Fluorescence , Lung/physiology , Microtubules , Mitosis/drug effects , Mitotic Index , Nocodazole/pharmacology , Resting Phase, Cell Cycle , Spindle ApparatusABSTRACT
The RAD51 protein has been shown to participate in homologous recombination by promoting ATP-dependent homologous pairing and strand transfer reactions. In the present study, we have investigated the possible involvement of RAD51 in non-homologous recombination. We demonstrate that overexpression of CgRAD51 enhances the frequency of spontaneous non-homologous recombination in the hprt gene of Chinese hamster cells. However, the rate of non-homologous recombination induced by the topoisomerase inhibitors campothecin and etoposide was not altered by overexpression of RAD51. These results indicate that the RAD51 protein may perform a function in connection with spontaneous non-homologous recombination that is not essential to or not rate-limiting for non-homologous recombination induced by camptothecin or etoposide. We discuss the possibility that the role played by RAD51 in non-homologous recombination observed here may not be linked to non-homologous end-joining.
Subject(s)
DNA-Binding Proteins/physiology , Enzyme Inhibitors/pharmacology , Recombination, Genetic/drug effects , Topoisomerase I Inhibitors , Animals , Camptothecin/pharmacology , Cell Line , Cricetinae , Cricetulus , DNA Topoisomerases, Type I/metabolism , DNA, Recombinant , DNA-Binding Proteins/genetics , Dose-Response Relationship, Drug , Etoposide/pharmacology , Gene Expression , Hypoxanthine Phosphoribosyltransferase/drug effects , Hypoxanthine Phosphoribosyltransferase/genetics , Hypoxanthine Phosphoribosyltransferase/metabolism , Plasmids/genetics , Point Mutation , Rad51 Recombinase , Recombination, Genetic/genetics , Transfection , Tumor Suppressor Protein p53/geneticsABSTRACT
The purpose of the descriptive study was to determine the hydration status of recreational backpackers (n = 201) hiking at altitudes between 7,500 and 14,000 feet. Urine specific gravity was used to document the level of hydration of each subject entering or leaving the Bridger-Teton Wilderness. Demographic, risk, and knowledge factors were also obtained from the sample. Both pre-hike and post-hike subjects were dehydrated; pre-hike mean specific gravity was 1.018, and the post-hike mean was 1.023, showing a significant difference (t = -4.671, p < 0.0000). A small subset group (n = 10) entered both pre- and post-hike data and the findings were similar to the large group, showing a significant increase in specific gravity post-hike (t = -4.881, p < 0.0009). Interestingly, 24% (n = 130) of the post-hike males presented with hematuria.
