scholarly journals Centrosomes Split in the Presence of Impaired DNA Integrity during Mitosis

2003 ◽  
Vol 14 (5) ◽  
pp. 1993-2004 ◽  
Author(s):  
Henderika M.J. Hut ◽  
Willy Lemstra ◽  
Engbert H. Blaauw ◽  
Gert W.A. van Cappellen ◽  
Harm H. Kampinga ◽  
...  
Keyword(s):  
Dna Damage ◽  
Chinese Hamster Ovary ◽  
Mammalian Cells ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Dna Integrity ◽  
Ovary Cells ◽  
Daughter Cells ◽  
Multipolar Spindles ◽  
Drosophila Embryos

A well-established function of centrosomes is their role in accomplishing a successful mitosis that gives rise to a pair of identical daughter cells. We recently showed that DNA replication defects and DNA damage in Drosophila embryos trigger centrosomal changes, but it remained unclear whether comparable centrosomal responses can be provoked in somatic mammalian cells. To investigate the centrosomal organization in the presence of impaired DNA integrity, live and ultrastructural analysis was performed on γ-tubulin–GFP and EGFP–α-tubulin–expressing Chinese hamster ovary cells. We have shown that during mitosis in the presence of incompletely replicated or damaged DNA, centrosomes split into fractions containing only one centriole. This results in the formation of multipolar spindles with extra centrosome-like structures. Despite the extra centrosomes and the multipolarity of the spindles, cells do exit from mitosis, resulting in severe division errors. Our data provide evidence of a novel mechanism showing how numerous centrosomes and spindle defects can arise and how this can lead to the formation of aneuploid cells.

scholarly journals A fluorescent lipid analogue can be used to monitor secretory activity and for isolation of mammalian secretion mutants.

1995 ◽  
Vol 6 (2) ◽  
pp. 135-150 ◽  
Author(s):  
N T Ktistakis ◽  
C Y Kao ◽  
R H Wang ◽  
M G Roth
Keyword(s):  
Chinese Hamster Ovary ◽  
Mammalian Cells ◽  
Secretory Pathway ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Brefeldin A ◽  
Secretory Activity ◽  
Specific Marker ◽  
Ovary Cells ◽  
Fluorescent Lipid

The use of reporter proteins to study the regulation of secretion has often been complicated by posttranslational processing events that influence the secretion of certain proteins, but are not part of the cellular mechanisms that specifically regulate secretion. This has been a particular limitation for the isolation of mammalian secretion mutants, which has typically been a slow process. To provide a reporter of secretory activity independent of protein processing events, cells were labeled with the fluorescent lipid analogue C5-DMB-ceramide (ceramide coupled to the fluorophore boron dipyrromethene difluoride) and its secretion was followed by fluorescence microscopy and fluorescence-activated cell sorting. Brefeldin A, which severely inhibits secretion in Chinese hamster ovary cells, blocked secretion of C5-DMB-ceramide. At high temperature, export of C5-DMB-ceramide was inhibited in HRP-1 cells, which have a conditional defect in secretion. Using C5-DMB-ceramide as a reporter of secretory activity, several different pulse-chase protocols were designed that selected mutant Chinese hamster ovary cells that were resistant to the drug brefeldin A and others that were defective in the transport of glycoproteins to the cell surface. Mutant cells of either type were identified in a mutagenized population at a frequency of 10(-6). Thus, the fluorescent lipid C5-DMB-ceramide can be used as a specific marker of secretory activity, providing an efficient, general approach for isolating mammalian cells with defects in the secretory pathway.

Choline Chloride Improves the Desiccation Tolerance of Chinese Hamster Ovary Cells

2010 ◽  
Author(s):  
Nilay Chakraborty ◽  
Michael A. Menze ◽  
Heidi Elmoazzen ◽  
Steve C. Hand ◽  
Mehmet Toner
Keyword(s):  
Chinese Hamster Ovary ◽  
Desiccation Tolerance ◽  
Mammalian Cells ◽  
Cell Injury ◽  
Chinese Hamster Ovary Cells ◽  
Choline Chloride ◽  
Chinese Hamster ◽  
Ovary Cells ◽  
Sodium Toxicity ◽  
Ionic Imbalance

Recently there has been much interest in using sugars such as trehalose to preserve mammalian cells in a dry state as an alternative to cryopreservation (1–5). However, some studies indicate that sugars alone may not be sufficient to prevent cell injury during drying. Other factors like sodium toxicity, ionic imbalance and pH excursions during dehydration are a few of the mechanisms that have been hypothesized to decrease the viability of mammalian cells. In the present study, we investigated whether or not substituting sodium chloride with choline chloride (2-hydroxy-N, N,N-trimethylethanaminium chloride) in the preservation medium improves desiccation tolerance of Chinese Hamster Ovary (CHO) cells.

scholarly journals Fluoride does not induce DNA breakage in Chinese hamster ovary cells in vitro

2004 ◽  
Vol 18 (3) ◽  
pp. 192-196 ◽  
Author(s):  
Daniel Araki Ribeiro ◽  
Clarissa Scolastici ◽  
Mariângela Esther Alencar Marques ◽  
Daisy Maria Fávero Salvadori
Keyword(s):  
Dna Damage ◽  
Chinese Hamster Ovary ◽  
Chinese Hamster Ovary Cells ◽  
Genetic Material ◽  
Chinese Hamster ◽  
Potential Health ◽  
Ovary Cells ◽  
The Mean ◽  
Genotoxicity Tests

Fluoride has been widely used in dentistry because it is a specific and effective caries prophylactic agent. However, excess fluoride may represent a hazard to human health, especially by causing injury to genetic material. Genotoxicity tests represent an important part of cancer research to assess the risk of potential carcinogens. In the current study, the potential DNA damage associated with exposure to fluoride was assessed by the single cell gel (comet) assay in vitro. Chinese hamster ovary cells were exposed to sodium fluoride (NaF) at final concentration ranging from 7 to 100 µg/ml for 3 h, at 37°C. The results pointed out that NaF in all concentrations tested did not contribute to DNA damage as depicted by the mean tail moment and tail intensity. These findings are clinically important since they represent an important contribution to a correct evaluation of the potential health risk associated with the exposure to dental agents.

Purification of phosphatidylglycerophosphate synthase from Chinese hamster ovary cells

2001 ◽  
Vol 354 (1) ◽  
pp. 9-15 ◽  
Author(s):  
Kiyoshi KAWASAKI ◽  
Osamu KUGE ◽  
Yoshio YAMAKAWA ◽  
Masahiro NISHIJIMA
Keyword(s):  
Chinese Hamster Ovary ◽  
Mammalian Cells ◽  
Polyclonal Antibodies ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Mitochondrial Fraction ◽  
Ovary Cells ◽  
Protein Levels ◽  
Defective Mutant ◽  
Sds Page

Phosphatidylglycerophosphate (PGP) synthase catalyses the committed step in the biosynthesis of phosphatidylglycerol and cardiolipin in mammalian cells. Recently we isolated a Chinese hamster ovary (CHO) PGS1 cDNA encoding PGP synthase. In the present study we purified this PGP synthase to near-homogeneity from the mitochondrial fraction of CHO-K1 cells; the final enzyme preparation gave a single 60kDa protein on SDS/PAGE. Polyclonal antibodies raised against a recombinant CHO PGS1 protein cross-reacted with the purified 60kDa protein and with CHO membrane proteins of 60kDa and 62kDa that increased after transfection with the PGS1 cDNA. The 60 and 62kDa protein levels in a PGP synthase-defective mutant of CHO-K1 cells were markedly lower than those in CHO-K1 cells. These results indicated that the purified 60kDa protein was PGP synthase encoded by the PGS1 gene. In addition we found that the purified PGP synthase had no PGP phosphatase activity, indicating that phosphatidylglycerol was produced from CDP-diacylglycerol through two steps catalysed by distinct enzymes, PGP synthase and PGP phosphatase.

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