Oncogenic transformation of murine C3H 10T1/2 cells resulting from DNA double-strand breaks induced by a restriction endonuclease

Pe, Bryant; Riches, A.C.

doi:10.1038/bjc.1989.378

Cited by 35 publications

(13 citation statements)

References 15 publications

(19 reference statements)

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“…Alteration of cell morphology and growth pattern in the M. fermentans-infected cultures was noted after passage 6. Similar changes were also found in M. penetrans-infected cultures, but only after passage [15][16]. In comparison, none of the mycoplasma-free control cultures passed in parallel showed morphological transformation.…”

Section: Resultssupporting

confidence: 57%

“…Since the question of whether mycoplasmas can induce malignant transformation of mammalian cells may have great significance in general biology, tumor biology, as well as direct clinical implications, we have examined mycoplasmal transforming effects in the murine embryonic C3H/10T½/2 (C3H) cell system, pne of the few standard test systems available for studying potential carcinogenic agents or factors in animals or humans (12)(13)(14)(15). Using this model system with low inherent spontaneous transformation (12), we did not find mycoplasmas induced acute mammalian cell transformation described in the earlier studies.…”

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confidence: 99%

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Mycoplasmas and oncogenesis: persistent infection and multistage malignant transformation.

Tsai

Wear

Shih

et al. 1995

Proc. Natl. Acad. Sci. U.S.A.

169

171

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Oncogenic potential of human mycoplasmas was studied using cultured mouse embryo cells, C3H/10T1/2 (C3H). Mycoplasma fermentans and Mycoplasma penetrans, mycoplasmas found in unusually high frequencies among patients with AIDS, were examined. Instead of acute transformation, a multistage process in promotion and progression of malignant cell transformation with long latency was noted; after 6 passages (1 wk per passage) of persistent infection with M. fermentans, C3H cells exhibited phenotypic changes with malignant characteristics that became progressively more prominent with further prolonged infection. Up to at least the 11th passage, all malignant changes were reversible if mycoplasmas were eradicated by antibiotic treatment. Mycoplasma fermentans and Mycoplasma penetrans are the most common mycoplasmas associated with AIDS (1-3). M penetrans infects male homosexuals and is seroepidemiologically associated with Kaposi sarcoma (4). Wall-free mycoplasmas are among the few prokaryotes that can grow symbiotically and have a close interaction with mammalian cells for long periods of time without causing acute cytopathic effects. Thus, infections in cell culture are commonly unrecognized, and many persistent parasitic infections in humans or in animals are clinically silent (5,6). But what is the potential effect on mammalian hosts parasitically infected by the prokaryotes with seemingly low virulence? Can mycoplasmas, the simplest organisms capable of self-replication, alter significantly and permanently the biological properties of mammalian cells during the long course of persistent infection?In the 1960s, two studies reported that infection of mycoplasmas (Mycoplasma orale and one unspeciated) caused chromosomal changes (7,8). But the studies did not reveal that the cells underwent transformation. A separate study reported that introduction of mycoplasmas into cultures of baby hamster kidney (BHK) cells produced immediate morphological transformation with a higher soft agar cloning efficiency (9). Unfortunately, the BHK cells had a high rate of spontaneous transformation. In the 1980s, one article reported that an arthropod spiroplasma could rapidly transform mouse and monkey cells (10). Most scientists thought mycoplasmas simply introduced confusing artifacts that mimic cell transformation (11).Since the question of whether mycoplasmas can induce malignant transformation of mammalian cells may have great significance in general biology, tumor biology, as well as direct clinical implications, we have examined mycoplasmal transforming effects in the murine embryonic C3H/10T½/2 (C3H) cell system, pne of the few standard test systems available for studying potential carcinogenic agents or factors in animals or humans (12)(13)(14)(15). Using this model system with low inherent spontaneous transformation (12), we did not find mycoplasmas induced acute mammalian cell transformation described in the earlier studies. Instead, we found mycoplasma-mediated oncogenesis had a long latency and required a chronic pe...

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Section: Resultssupporting

confidence: 57%

mentioning

confidence: 99%

Mycoplasmas and oncogenesis: persistent infection and multistage malignant transformation.

Tsai

Wear

Shih

et al. 1995

Proc. Natl. Acad. Sci. U.S.A.

169

171

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“…When genomic DNA strand breaks have been generated by the introduction of restriction endonucleases into cells, a strikingly increased frequency of chromosomal aberrations has been observed in cells reaching mitosis (6,7). In addition, restriction enzymeinduced strand breaks have been found to increase neoplastic transformation (5,8,68). Finally, unrepaired or irreparable DNA strand breaks may trigger cell death.…”

Section: Discussionmentioning

confidence: 99%

DNA strand breaks: the DNA template alterations that trigger p53-dependent DNA damage response pathways.

Nelson¹,

Kastan²

1994

Mol. Cell. Biol.

741

344

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The tumor suppressor protein p53 serves as a critical regulator of a G1 cell cycle checkpoint and of apoptosis following exposure of cells to DNA-damaging agents. The mechanism by which DNA-damaging agents elevate p53 protein levels to trigger G1/S arrest or cell death remains to be elucidated. In fact, whether damage to the DNA template itself participates in transducing the signal leading to p53 induction has not yet been demonstrated. We exposed human cell lines containing wild-type p53 alleles to several different DNA-damaging agents and found that agents which rapidly induce DNA strand breaks, such as ionizing radiation, bleomycin, and DNA topoisomerase-targeted drugs, rapidly triggered p53 protein elevations. In addition, we determined that camptothecin-stimulated trapping of topoisomerase I-DNA complexes was not sufficient to elevate p53 protein levels; rather, replication-associated DNA strand breaks were required. Furthermore, treatment of cells with the antimetabolite N(phosphonoacetyl)-L-aspartate (PALA) did not cause rapid p53 protein increases but resulted in delayed increases in p53 protein levels temporally correlated with the appearance of DNA strand breaks. Finally, we concluded that DNA strand breaks were sufficient for initiating p53-dependent signal transduction after finding that introduction of nucleases into cells by electroporation stimulated rapid p53 protein elevations. While DNA strand breaks appeared to be capable of triggering p53 induction, DNA lesions other than strand breaks did not. Exposure of normal cells and excision repair-deficient xeroderma pigmentosum cells to low doses of UV light, under conditions in which thymine dimers appear but DNA replication-associated strand breaks were prevented, resulted in p53 induction attributable to DNA strand breaks associated with excision repair. Our data indicate that DNA strand breaks are sufficient and probably necessary for p53 induction in cells with wild-type p53 alleles exposed to DNA-damaging agents.

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“…DNA double-strand breaks can cause chromosomal rearrangements (1), which can lead to cell killing (13), mutagenesis (25), and cell transformation (5). To understand how DNA double-strand breaks lead to genetic rearrangements, it is important to understand the mechanisms of DNA double-strand-break rejoining and how these processes are carried out at the DNA and chromosomal levels.…”

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confidence: 99%

Illegitimate recombination induced by DNA double-strand breaks in a mammalian chromosome.

Phillips¹,

Morgan²

1994

Mol. Cell. Biol.

110

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We examined DNA double-strand-break-induced mutations in the endogenous adenine phosphoribosyltransferase (APRT) gene in cultured Chinese hamster ovary cells after exposure to restriction endonucleases. PvuII, EcoRV, and StuI, all of which produce blunt-end DNA double-strand breaks, were electroporated into CHO-AT3-2 cells hemizygous at the APRT locus. Colonies of viable cells containing mutations at APRT were expanded, and the mutations that occurred during break repair were analyzed at the DNA sequence level. Restriction enzyme-induced mutations consisted of small deletions of 1 to 36 bp, insertions, and combinations of insertions and deletions at the cleavage sites. Most of the small deletions involved overlaps of one to four complementary bases at the recombination junctions. Southern blot analysis revealed more complex mutations, suggesting translocation, inversion, or insertion of larger chromosomal fragments. These results indicate that blunt-end DNA double-strand breaks can induce illegitimate (nonhomologous) recombination in mammalian chromosomes and that they play an important role in mutagenesis.In the mammalian genome, DNA double-strand breaks can occur during cellular processes (4,46) or as the result of exposure to DNA-damaging agents (51). DNA double-strand breaks can cause chromosomal rearrangements (1), which can lead to cell killing (13), mutagenesis (25), and cell transformation (5). To understand how DNA double-strand breaks lead to genetic rearrangements, it is important to understand the mechanisms of DNA double-strand-break rejoining and how these processes are carried out at the DNA and chromosomal levels.Studies of plasmid integration in a variety of mammalian cell types, along with simian virus 40 (SV40) recircularization studies with monkey cells, have shown that mammalian cells predominantly repair DNA double-strand breaks by endjoining mechanisms that do not require extensive homology between the molecules to be joined (39). Recombination between nonhomologous DNA substrates was first described for bacteria (15) and has been defined as illegitimate recombination (16). Most studies examining illegitimate recombination have used DNA substrates linearized with various restriction endonucleases to produce specific combinations of end structures. These substrates are cleaved in vitro and passed through various cellular and cell-free systems, and the rejoined products are studied. Recircularization studies of a linearized SV40 genome passed through CV 1 monkey cells, and plasmid rejoining studies with Xenopus laevis egg and human cell extracts, have shown that these end-joining processes frequently use overlaps of one to six complementary bases (36,44,48). The products of illegitimate recombination between segments of mammalian chromosomes have been studied by examination of translocation breakpoints and deletion and insertion junctions at various genetic loci (24,25 bination, the initial steps leading to the recombinational event and the mechanisms involved are not understood.To study the...

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Oncogenic transformation of murine C3H 10T1/2 cells resulting from DNA double-strand breaks induced by a restriction endonuclease

Cited by 35 publications

References 15 publications

Mycoplasmas and oncogenesis: persistent infection and multistage malignant transformation.

Mycoplasmas and oncogenesis: persistent infection and multistage malignant transformation.

DNA strand breaks: the DNA template alterations that trigger p53-dependent DNA damage response pathways.

Illegitimate recombination induced by DNA double-strand breaks in a mammalian chromosome.

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