Inactivation of Cdc2 increases the level of apoptosis induced by DNA damage

1995 ◽  
Vol 108 (8) ◽  
pp. 2897-2904 ◽  
Author(s):  
W. Ongkeko ◽  
D.J. Ferguson ◽  
A.L. Harris ◽  
C. Norbury
Keyword(s):  
Dna Damage ◽  
Cell Death ◽  
Protein Kinase ◽  
Apoptotic Cell ◽  
Mutant Cell ◽  
Strand Break ◽  
Temperature Sensitive ◽  
Dna Strand Break ◽  
Dna Strand ◽  
Inhibitor Compounds

A number of lines of evidence have suggested a possible involvement of the mitosis-promoting protein kinase Cdc2 in the process of apoptotic cell death, and one recent study concluded that premature activation of Cdc2 is required for apoptosis. Here we have used a temperature-sensitive murine Cdc2 mutant cell line and Cdc2 inhibitor compounds to study the effect of inhibition of this protein kinase on apoptosis induced by DNA-damaging drugs. Inhibition of Cdc2 activity before or during exposure to DNA strand break-inducing drugs had the effect of increasing the level of subsequent apoptosis, as assessed by electron microscopy and flow cytometry. We conclude that, far from being required for cell death, a form of mammalian Cdc2 suppresses apoptosis induced by DNA damage. This form of Cdc2 appears to be active in G2-arrested cells and is therefore presumably distinct from the mitosis-promoting Cdc2-cyclin B heterodimer.

scholarly journals Natural Merosesquiterpenes Activate the DNA Damage Response via DNA Strand Break Formation and Trigger Apoptotic Cell Death in p53-Wild-type and Mutant Colorectal Cancer

Cancers ◽  
2021 ◽  
Vol 13 (13) ◽  
pp. 3282
Author(s):  
Apisada Jiso ◽  
Philipp Demuth ◽  
Madeleine Bachowsky ◽  
Manuel Haas ◽  
Nina Seiwert ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Dna Damage ◽  
Cell Death ◽  
Dna Damage Response ◽  
Strand Break ◽  
Wild Type ◽  
Dna Strand Break ◽  
Damage Response ◽  
Dna Strand ◽  
Break Formation

Colorectal cancer (CRC) is a frequently occurring malignant disease with still low survival rates, highlighting the need for novel therapeutics. Merosesquiterpenes are secondary metabolites from marine sponges, which might be useful as antitumor agents. To address this issue, we made use of a compound library comprising 11 isolated merosesquiterpenes. The most cytotoxic compounds were smenospongine > ilimaquinone ≈ dactylospontriol, as shown in different human CRC cell lines. Alkaline Comet assays and γH2AX immunofluorescence microscopy demonstrated DNA strand break formation in CRC cells. Western blot analysis revealed an activation of the DNA damage response with CHK1 phosphorylation, stabilization of p53 and p21, which occurred both in CRC cells with p53 knockout and in p53-mutated CRC cells. This resulted in cell cycle arrest followed by a strong increase in the subG1 population, indicative of apoptosis, and typical morphological alterations. In consistency, cell death measurements showed apoptosis following exposure to merosesquiterpenes. Gene expression studies and analysis of caspase cleavage revealed mitochondrial apoptosis via BAX, BIM, and caspase-9 as the main cell death pathway. Interestingly, the compounds were equally effective in p53-wild-type and p53-mutant CRC cells. Finally, the cytotoxic activity of the merosesquiterpenes was corroborated in intestinal tumor organoids, emphasizing their potential for CRC chemotherapy.

scholarly journals DNA damage-induced cell cycle checkpoints and DNA strand break repair in development and tumorigenesis

Oncogene ◽  
1999 ◽  
Vol 18 (55) ◽  
pp. 7883-7899 ◽  
Author(s):  
Gopal K Dasika ◽  
Suh-Chin J Lin ◽  
Song Zhao ◽  
Patrick Sung ◽  
Alan Tomkinson ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Strand Break ◽  
Cell Cycle Checkpoints ◽  
Dna Strand Break ◽  
Break Repair ◽  
Dna Strand

Evaluation of genotoxic potential of commonly used organophosphate pesticides in peripheral blood lymphocytes of rats

2014 ◽  
Vol 34 (4) ◽  
pp. 390-400 ◽  
Author(s):  
A Ojha ◽  
YK Gupta
Keyword(s):  
Dna Damage ◽  
Damage Index ◽  
Fluorescence Emission ◽  
Strand Break ◽  
Peripheral Blood Lymphocytes ◽  
Dependent Manner ◽  
Strand Breaks ◽  
Dna Strand Break ◽  
Cross Links ◽  
Dna Strand

Chlorpyrifos (CPF), methyl parathion (MPT), and malathion (MLT) are among the most extensively used organophosphate (OP) pesticides in India. DNA protein cross-links (DPC) and DNA strand breaks are toxic lesions associated with the mechanism(s) of toxicity of carcinogenic compounds. In the present study, we examined the hypothesis that individual and interactive genotoxic effects of CPF, MPT, and MLT are involved in the formation of DPC and DNA strand break. The DNA strand break was measured by comet assay and expressed as DNA damage index, while DPC estimation was carried out by fluorescence emission assay. The results showed that exposure of rat lymphocytes with CPF, MPT, and MLT caused significantly marked increase in DNA damage and DPC formation in time-dependent manner. MPT caused the highest damage, and these pesticides do not potentiate the toxicity of each other.

Specific transcripts are elevated in Saccharomyces cerevisiae in response to DNA damage

1984 ◽  
Vol 4 (11) ◽  
pp. 2356-2363
Author(s):  
T McClanahan ◽  
K McEntee
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Damage ◽  
Southern Hybridization ◽  
Single Copy ◽  
Strand Break ◽  
Yeast Genome ◽  
Dna Strand Break ◽  
Isolation And Characterization ◽  
Dna Strand

Differential hybridization has been used to identify genes in Saccharomyces cerevisiae displaying increased transcript levels after treatment of cells with UV irradiation or with the mutagen/carcinogen 4-nitroquinoline-1-oxide (NQO). We describe the isolation and characterization of four DNA damage responsive genes obtained from screening ca. 9,000 yeast genomic clones. Two of these clones, lambda 78A and pBR178C, contain repetitive elements in the yeast genome as shown by Southern hybridization analysis. Although the genomic hybridization pattern is distinct for each of these two clones, both of these sequences hybridize to large polyadenylated transcripts ca. 5 kilobases in length. Two other DNA damage responsive sequences, pBRA2 and pBR3016B, are single-copy genes and hybridize to 0.5- and 3.2-kilobase transcripts, respectively. Kinetic analysis of the 0.5-kilobase transcript homologous to pBRA2 indicates that the level of this RNA increases more than 15-fold within 20 min after exposure to 4-nitroquinoline-1-oxide. Moreover, the level of this transcript is significantly elevated in cells containing the rad52-1 mutation which are deficient in DNA strand break repair and gene conversion. These results provide some of the first evidence that DNA damage stimulates transcription of specific genes in eucaryotic cells.

scholarly journals Involvement of a periplasmic protein kinase in DNA strand break repair and homologous recombination in Escherichia coli

2007 ◽  
Vol 65 (6) ◽  
pp. 1595-1595
Author(s):  
Nivedita P. Khairnar ◽  
Vidya A. Kamble ◽  
Suhas H. Mangoli ◽  
Shree K. Apte ◽  
Hari S. Misra
Keyword(s):  
Escherichia Coli ◽  
Protein Kinase ◽  
Homologous Recombination ◽  
Strand Break ◽  
Periplasmic Protein ◽  
Dna Strand Break ◽  
Break Repair ◽  
Dna Strand

scholarly journals Protein kinase Cδ regulates vaccinia-related kinase 1 in DNA damage–induced apoptosis

2011 ◽  
Vol 22 (8) ◽  
pp. 1398-1408 ◽  
Author(s):  
Choon-Ho Park ◽  
Bo-Hwa Choi ◽  
Min-Woo Jeong ◽  
Sangjune Kim ◽  
Wanil Kim ◽  
...  
Keyword(s):  
Dna Damage ◽  
Cell Death ◽  
Protein Kinase ◽  
Cell Viability ◽  
Apoptotic Cell ◽  
Critical Role ◽  
Regulatory Region ◽  
Dependent Manner ◽  
Protein Kinase Cδ

Vaccinia-related kinase 1 (VRK1) is a novel serine/threonine kinase that plays an important role in cell proliferation. However, little is known about the upstream regulators of VRK1 activity. Here we provide evidence for a role of protein kinase Cδ (PKCδ) in the regulation of murine VRK1. We show that PKCδ interacts with VRK1, phosphorylates the Ser-355 residue in the putative regulatory region, and negatively regulates its kinase activity in vitro. Intriguingly, PKCδ-induced cell death was facilitated by phosphorylation of VRK1 when cells were exposed to a DNA-damaging agent. In addition, p53 played a critical role in the regulation of DNA damage–induced cell death accompanied by PKCδ-mediated modulation of VRK1. In p53-deficient cells, PKCδ-mediated phosphorylation of VRK1 had no effect on cell viability. However, cells overexpressing p53 exhibited significant reduction of cell viability when cotransfected with both VRK1 and PKCδ. Taken together, these results indicate that PKCδ regulates phosphorylation and down-regulation of VRK1, thereby contributing to cell cycle arrest and apoptotic cell death in a p53-dependent manner.

scholarly journals Involvement of a periplasmic protein kinase in DNA strand break repair and homologous recombination in Escherichia coli

2007 ◽  
Vol 65 (2) ◽  
pp. 294-304 ◽  
Author(s):  
Nivedita P. Khairnar ◽  
Vidya A. Kamble ◽  
Suhas H. Mangoli ◽  
Shree K. Apte ◽  
Hari S. Misra
Keyword(s):  
Escherichia Coli ◽  
Protein Kinase ◽  
Homologous Recombination ◽  
Strand Break ◽  
Periplasmic Protein ◽  
Dna Strand Break ◽  
Break Repair ◽  
Dna Strand

scholarly journals The Enzyme-Modified Neutral Comet (EMNC) Assay for Complex DNA Damage Detection

2021 ◽  
Vol 4 (1) ◽  
pp. 14
Author(s):  
Maria Rita Fabbrizi ◽  
Jonathan R. Hughes ◽  
Jason L. Parsons
Keyword(s):  
Dna Damage ◽  
Complex Analysis ◽  
Single Cells ◽  
Strand Break ◽  
Dna Lesions ◽  
Repair Capacity ◽  
Strand Breaks ◽  
Dna Strand Break ◽  
Repair Enzymes ◽  
Dna Strand

The comet assay is a versatile, simple, and sensitive gel electrophoresis–based method that can be used to measure and accurately quantify DNA damage, particularly single and double DNA strand breaks, in single cells. While generally this is used to measure variation in DNA strand break levels and repair capacity within a population of cells, the technique has more recently been adapted and evolved into more complex analysis and detection of specific DNA lesions, such as oxidized purines and pyrimidines, achieved through the utilization of damage-specific DNA repair enzymes following cell lysis. Here, we detail a version of the enzyme-modified neutral comet (EMNC) assay for the specific detection of complex DNA damage (CDD), defined as two or more DNA damage lesions within 1–2 helical turns of the DNA. CDD induction is specifically relevant to ionizing radiation (IR), particularly of increasing linear energy transfer (LET), and is known to contribute to the cell-killing effects of IR due to the difficult nature of its repair. Consequently, the EMNC assay reveals important details regarding the extent and complexity of DNA damage induced by IR, but also has potential for the study of other genotoxic agents that may induce CDD.

scholarly journals Defective DNA Strand Break Repair after DNA Damage in Prostate Cancer Cells

2004 ◽  
Vol 64 (23) ◽  
pp. 8526-8533 ◽  
Author(s):  
Rong Fan ◽  
Tirukalikundram S. Kumaravel ◽  
Farid Jalali ◽  
Paula Marrano ◽  
Jeremy A. Squire ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Dna Damage ◽  
Cancer Cells ◽  
Strand Break ◽  
Prostate Cancer Cells ◽  
Dna Strand Break ◽  
Break Repair ◽  
Dna Strand
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