Mutations in the p53 gene in radiation-sensitive and -resistant human squamous carcinoma cells (original) (raw)
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Mutations in the p53 Gene in Radiation-sensitive and -resistant Human Squamous Carcinoma Cells1
1992
Five of six human squamous cell carcinoma (SCC) cell lines charac terized as radiation sensitive (SQ-38, SCC-9, SQ-9G) or radiation re sistant (SQ-20B, SCC-35, JSQ-3) exhibited alterations of the p53 gene. The point mutations and a deletion were detected by using single- stranded conformational polymorphism analysis and polymerase chain reaction-direct sequencing. Interestingly, three of three radiation-sensi tive and two of
The Journal of Pathology, 2003
Although little is understood of the underlying mechanisms, there are tissue-specific responses to tumourigenic and therapeutic agents and these responses are influenced by genetic factors. Ionizing radiation is an important tumourigenic and therapeutic agent for which there is substantial evidence for such tissue-dependent and genotype-dependent responses. Because the p53 tumour suppressor protein is a major determinant of cellular responses to radiation, the present study has investigated whether modification of the p53 pathway contributes to tissue-dependent and genotype-dependent responses using inbred strains of mice. Comparison of responses in haemopoietic and epithelial cells in irradiated C57BL/6 and DBA/2 mice revealed significant differences in p53 and apoptotic responses in different cell types and in different cells of the same type, reflecting the complexity of damage responses operating in the whole organism. The data suggest that p53-mediated up-regulation of Bax is a major determinant of apoptosis in the spleen, but not in the intestine, whereas p53-mediated induction of p21 waf1 plays an anti-apoptotic role in the spleen, but not in the intestine. It is also shown that p53 stabilization and differential transactivational activities towards Bax or p21 waf1 are influenced by genetic factors that act in a tissue-specific manner. Analysis of ATM, a potential mediator of differential p53 activation, indicates that this key regulator of radiation responses is preferentially induced in epithelial cells, but is unlikely to account for genetic modification of p53 or apoptotic responses in the mouse strains studied. Polymorphisms in the p53 or DNA-PKcs genes are also unlikely to account for the genetic modifications that are reported here. There are numerous further potential modifiers of the p53 pathway, but analysis of backcross and inter-cross mice demonstrates that genes responsible for the complex modification of these in vivo responses can be identified by linkage analysis. This approach has the potential to reveal new or unexpected interactions involving the p53 pathway that determine both shortterm and long-term effects of radiation exposure and the basis of tissue-specific responses and tumour susceptibility.
Relationship between p53 status and radiosensitivity in human tumour cell lines
British journal of cancer, 1996
We examined the relationship between p53 levels before and after irradiation, radiation-induced cell cycle delays, apoptotic cell death and radiosensitivity in a panel of eight human tumour cell lines. The cell lines differed widely in their clonogenic survival after radiation, (surviving fraction at 2 Gy: SF2=0.18-0.82). Constitutive p53 protein levels varied from 2.2 +/- 0.4 to 6.3 +/- 0.3 optical density units (OD) per 10(6) cells. p53 after irradiation (6 Gy) also varied between the cell lines, ranging from no induction to a 1.6-fold increase in p53 levels 4 h after treatment. p53 function was also assessed by G1 cell cycle arrest after irradiation. The cellular response to radiation, measured as G0/G1 arrest, and the induction of apoptosis were in good agreement. However, a trace amount of DNA ladder formation was found in two cell lines lacking G1 arrest. Overall cellular radiosensitivity correlated well with the level of radiation-induced G1 arrest (correlation coefficient r=...
Stem Cells, 1995
We investigated the possible relationship between immunohistochemically detected p53 expression and in vitro response to γ-irradiation in 24 primary cultures of human ovarian cancers and cutaneous melanomas. The frequency of p53-positive tumors was around 60% within each tumor histotype. The range of the surviving fraction at 2 Gy (SF2) was similar in p53-positive (0.10-0.76) and p53-negative (0.23-0.65) tumors, with median values of 0.36 and 0.33, respectively. No differences were observed in the accumulation of DNA-double strand breaks, assessed by neutral filter elution after exposure to 50 Gy, between p53-positive and p53-negative tumors. As regards DNA lesion repair, after 2 h of recovery the percentage of rejoined DNA-double strand breaks ranged from 19% to 99% in the different cultures, but again the distribution of values was similar for p53-positive and p53-negative tumors. Specifically, the median percentage of repaired DNA-double strand breaks was 70% and 74% in the two groups. On the whole, our data do not support the hypothesis that p53 overexpression is a major determinant of in vitro radiation response.
Spontaneous and radiation-induced tumorigenesis in p53-deficient mice
International Congress Series, 2002
To investigate the spontaneous and radiation induced tumorigenesis, tumor development and molecular changes were examined in p53 gene-deficient mice. All of the null (p53 À / À ) animals died of leukemia, sarcoma, etc. before 12 months of age, and predominantly of lymphocytic leukemia at the early age of 3 -5 months, while heterozygous (p53 +/ À ) and wild type (p53 +/ + ) animals survived more than 30 and 40 months, respectively. Half of the heterozygotes developed sarcomas (osteosarcomas, rhabdomyosarcomas, hemangiomas), and some developed tumors in the lung and liver; however, a lower incidence of leukemia was found. The onset of the tumor was late in their lives in the heterozygotes, though the tumors grew rapidly. In the wild type mice, leukemia and sarcomas were rarely observed. The 137 Cs g-irradiation shortened the lifespan of the heterozygotes with an earlier tumor development, though this effect was not evident in the null mice. Among the 54 spontaneous tumors studied in the heterozygotes, 12 (22.2%) showed loss of the wild type p53 allele (loss of heterozygosity, LOH) and one functional mutation was found at exon 6 of the p53 gene. All the LOH was found in leukemia and sarcomas, except in one breast carcinoma. D
Cancer research, 1994
The present study assessed the role ofthep53 tumor suppressor gene in cell cycle arrest and apoptosis following treatment of Burldtt's lymphoma and lymphoblastoid cell lines with ‘y-rays, etoposide, nitrogen mustard, and cisplatin. Cell cycle arrest was measured by flow cytometry; p5.3 and @ protein levels were measured by Western blotting; cell sur vival was measured in 72-96-h growth inhibition assays and by trypan blue staining, and apoptotic DNA fragmentation was assessed by either agarose gel electrophoresis or a modified ifiter elution method. We found that y-rays and etoposide induced a strong G1 arrest in the wild-typeps3 lines while nitrogen mustard and cisplatin induced relatively little G1 arrest. All agents failed to induce G1 arrest in cells containing mutantp53 genes. The degree of C1 arrest observed with these agents correlated with the rate ofp53 and p2JW41fI/C4PI protein accumulation: ‘y-rays and etopo side induced rapid accumulation of both p5.3 and p21@'@―1; nitrogen mustard and cisplatin induced slow accumulation of p5.3 and no major accumulation of the @2jWafI/C1@1 protein. Despite differences in G1 arrest and kinetics ofp53 orp21@―@ protein accumulation, all agents tended to decrease survival to a greater extent in the wild-type p5.3 lines com pared to the mutant p5.3 lines. Cell death in the wild-type p53 lines was associated with intracellular DNA degradation into oligonucleosomal sized DNA fragments, indicative ofapoptosis.
Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis, 1997
The p53 tumor suppressor protein plays an important role in regulating the cellular response to DNA damage, including cell cycle arrest and apoptosis induction. Normal p53 function is critical for the maintenance of genomic stability. The mouse lymphoma L5178YrTK qry -3.7.2C cell line is widely used in genetic toxicology for mutagenesis and clastogenesis testing. A related line L5178Y-R, has previously been shown to react with antibodies specific for mutant as well as wild-type p53 protein and to exhibit delayed cell death after radiation. For this reason, as well as the mouse lymphoma assay's reputation for high sensitivity of detection for genotoxic agents but low specificity, we examined several clones of Ž . L5178Y cells for mutations in the conserved core domain exons 5-8 of the p53 gene. Using single-strand conformational polymorphism analysis, we found evidence for the same mutation in exon 5 of p53 in L5178Y-R, L5178Y-S and qrq Ž . Ž .
Toxicologic Pathology, 2005
Treatment with 8-methoxypsoralen (8-MOP) and ultraviolet radiation (primarily UVA), called PUVA therapy, has been used to treat different chronic skin diseases but led to a significant increased risk for skin cancer. The National Toxicology Program (NTP) performed a study in mice treated with PUVA that showed a significant increase in squamous cell carcinomas of the skin. In the present study, we evaluated the protein expression of p53 and PCNA and DNA mutations of p53 and H-ras genes in both hyperplastic and neoplastic squamous cell lesions from the NTP study. By immunohistochemical staining, protein expression of both p53 and PCNA was detected in 3/16 (19%) of hyperplastic lesions and 14/17 (82%) of SCCs in groups treated with both 8-MOP and UVA. The mutation frequency of p53 in SCCs from mice administered 8-MOP plus UVA was 15/17 (88%) with a predominant distribution of mutations in exon 6 (14/15 -93%). No H-ras mutations were detected in the hyperplastic lesions/tumors. The mutagenic effect of PUVA on the p53 tumor suppressor gene may lead to a conformational modification and inactivation of the p53 protein, which are considered critical steps in PUVA-induced skin carcinogenesis. The p53 mutational frequency and patterns from our study were different from those reported in human PUVA-type tumors.
The 1993 Walter Hubert Lecture: the role of the p53 tumour-suppressor gene in tumorigenesis
1994
The p53 tumour-suppressor gene is mutated in 60% of human tumours, and the product of the gene acts as a suppressor of cell division. It is thought that the growth-suppressive effects of p53 are mediated through the transcriptional transactivation activity of the protein. Overexpression of the p53 protein results either in arrest in the GI phase of the cell cycle or in the induction of apoptosis. Both the level of the protein and its transcriptional transactivation activity increase following treatment of cells with agents that damage DNA, and it is thought that p53 acts to protect cells against the accumulation of mutations and subsequent conversion to a cancerous state. The induction of p53 levels in cells exposed to gamma-irradiation results in cell cycle arrest in some cells (fibroblasts) and apoptosis in others (thymocytes). Cells lacking p53 have lost this cell cycle control and presumably accumulate damage-induced mutations that result in tumorigenesis. Thus, the role of p53 in suppressing tumorigenesis may be to rescue the cell or organism from the mutagenic effects of DNA damage. Loss of p53 function accelerates the process of tumorigenesis and alters the response of cells to agents that damage DNA, indicating that successful strategies for radiation therapy may well need to take into account the tissue of origin and the status of p53 in the tumour.