Molecular mechanism of thymine-less death

Mennigmann, H. D.; Szybalski, Waclaw

doi:10.1016/0006-291x(62)90023-2

Cited by 72 publications

(22 citation statements)

References 5 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…3 G and H). It is of interest that DNA strand breaks in bacterial (11)(12)(13)(14)(15) and in mammalian cells (25) undergoing thymineless death appear not to be repaired when cells are supplemented with thymine or thymidine.…”

Section: Resultsmentioning

confidence: 99%

“…In addition, there was suggestive evidence that newly formed DNA may be partially degraded, perhaps as a result of misincorporation of dUMP and the subsequent repair reactions (7). Misincorporation of dUMP also has been demonstrated in cells treated with MTX (8), and it was hypothesized that futile repair processes could result in extensive DNA degradation, by analogy to thymineless death in bacteria (9)(10)(11)(12)(13)(14)(15).…”

mentioning

confidence: 99%

See 1 more Smart Citation

Accumulation of DNA strand breaks and methotrexate cytotoxicity.

Li¹,

Kaminskas²

1984

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

113

View full text Add to dashboard Cite

There was a progressive formation of strand breaks in mature DNA of Ehrlich ascites tumor cells that were treated with methotrexate. Cells were labeled with [14C~thyMi-dine before incubation with methotrexate, and DNA strand breaks were measured by alkaline and by neutral filter elution methods. DNA single-strand breaks accumulated in a linear fashion as a function of time during the first 10 hr of incubation with 2 ,uM methotrexate. Thereafter, the accumulation of DNA strand breaks deviated from linearity because of progressive cell death. The extent of DNA strand breakage in cells that had been incubated with methotrexate for 24 hr was as high as in cells that had been irradiated with 300 rads. DNA strand breaks persisted in cells that were incubated, after exposure to methotrexate, in medium containing thymidine, hypoxanthine, and nonessential amino acids, indicating that these strand breaks were poorly repaired. Cell death commenced after 10 hr of incubation with methotrexate and continued during the following 3-4 days. These findings suggest that cell death was due to a lethal accumulation of DNA strand breaks. The formation of DNA strand breaks is probably due to inefficient DNA repair, resulting from the inhibition of syntheses of thymidylate and of purine nucleotides. The accumulation of DNA strand breaks was minimal in growth-arrested cells, which are resistant to methotrexate toxicity.Cytotoxicity of the antifolate methotrexate (MTX) is generally thought to be related to the inhibition of DNA synthesis, although the precise mechanism of cell death is not known (1). Multiplying cells are sensitive to MTX, especially cells in the S phase of the cell cycle (2), while resting cells are not (2-4). MTX restricts the synthesis of thymidylate and of purine nucleotides by inhibiting dihydrofolate reductase and, to a lesser extent, thymidylate synthetase (1). Hence, MTX cytotoxicity has been postulated to occur both by "thymineless death" and "purineless death" mechanisms (5). DNA synthesis is inhibited rapidly in cells that are incubated with micromolar concentrations of MTX. In CCRF-CEM human lymphoblastoid cells, DNA synthetic rates decrease by 80% within 30 min and then remain stable for at least 6 hr (6). The newly synthesized DNA accumulates in 80S fragments and does mature to bulk-size DNA upon reversal of the MTX block, at least within the first hr of MTX treatment (6). An abnormal progression of DNA synthesis, with accumulation of low molecular weight DNA fragments, also was demonstrated with the antifolate metoprine (7). In addition, there was suggestive evidence that newly formed DNA may be partially degraded, perhaps as a result of misincorporation of dUMP and the subsequent repair reactions (7). Misincorporation of dUMP also has been demonstrated in cells treated with MTX (8), and it was hypothesized that futile repair processes could result in extensive DNA degradation, by analogy to thymineless death in bacteria (9-15). (16). Ehrlich ascites tumor cells were grown in continuous spinner cu...

show abstract

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

Accumulation of DNA strand breaks and methotrexate cytotoxicity.

Li¹,

Kaminskas²

1984

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

113

View full text Add to dashboard Cite

show abstract

“…The mechanism of thymineless death is not yet clear. However, numerous investigations (Maaloe & Hanawalt, 1961;Menningmann & Szybalski, 1962;Maaloe, 1963) have shown that death in the absence of thymine is due to disturbances in normal DNA replication, while biosynthesis of other cell components appears to be normal.…”

Section: Introductionmentioning

confidence: 97%

A genetical study of thymineless mutants ofE. coliK12

Alikhanian¹,

Iljina²,

Kaliaeva³

et al. 1966

Genet. Res.

View full text Add to dashboard Cite

The 150 independently isolatedthy−mutants ofE. coliK12 Hfr 3.OSO were studied genetically and phenotypically. Variants were found among the mutants in respect to the lag period of thymineless death, and temperature sensitivity. The latter correlates with mutations located at a specific site on the genetical map.Thethylocus is located between thecysandser/glygenes, and is a linear structure where 134thymutants are distributed over more than 17 sites. The site distribution of the mutants is not regular: about a half of them (62) are localized within one site and all these are temperature-sensitive.Two further genes involving utilization of thymine—tlrandtd—were found. Mutations oftlrlead to a reduced thymine requirement (0·5 μg./ml.instead of 20 μg./ml.). A mutation oftdresults in thymidine sensitivity.This latter character is expressed when thetd-sallele is transferred intoE. coliK12, prototrophic for thymine, by conjugation. Thymidine inhibition can be reversed by the addition of any riboside to the growth medium. Both genes map at the proximal end of the Hfr 3.OSO chromosome and are linked with thethrgene. The most probable gene order is:tlr-td-thr.The following results have been obtained from14C-thymine incorporation experiments with wild-type cells, as well as withthy−tlr+andthy−tlr−cells: (1) Wild-type cells incorporate exogenous thymine extremely poorly, but incorporate thymidine better. (2) Thethy−tlr+mutants are able to incorporate thymine only when high concentration are used, but can utilize a low concentration of thymidine. (3) Thethy−mutants are able to incorporate exogenous thymine as well as thymidine at low concentration. (4) Thetlrmutation is a thymine-specific one.

show abstract

“…The relationship, however, between UV, mitomycin and thymine-less deaths is not altogether clear, since some E. coli strains (B, B 5 ) are hyper-sensitive to UV, mitomycin and thymine-less death whereas other strains (K 12 uvr and rec-) exhibit hypersensitivity only to UV and mitomycin but not to thymine-less death. It was concluded that a single defect in DNA repair connot satisfactorily account for both hypersensitivity to thymine-less death (single-strand breaks in DNA; MENNIGMANN and SzYBALSKI, 1962) and ultrasensitivity to either UV or mitomycin (CUMMINGS and TAYLOR, 1966).Breakdown of DNA in mitomycin-exposed cells depends on the presence of magnesium ions, is temperature-dependent, and leads to the formation of mononucleotides and free bases SHIIO et al, 1962). Synthesis of new nucleases observed during mitomycin-effected induction of Jl phages reflects the activity of the induced phage genome, since in the nonlysogenic host no new A.-specific nucleases are induced KORN and WEISS-BACH, 1964) and since the nuclease-deficient Asus mutants fail to show an increase in nuclease activity at inducing doses of mitomycin (RADDING, 1964).…”

mentioning

confidence: 96%

Antibiotics

1967

View full text Add to dashboard Cite

AII rights, especially that of tnmslation into foreign languages, reserved. It is also forbidden to reproduce this book, either whole DedicationFor most areas of scientific pursuit, there is usually that rare investigator who has the imagination to conceive ideas, who has faith in his visions, and who has the ability to critically test his concepts in the laboratory. Almost invariably, this scientist also inspires younger men to enthusiastically enter into his research program. To him should go the accolades and the recognitions of the esteem in which he is held. As only a small part of this esteem, we wish to dedicate these books to Professor SELMAN A. W AKSMAN in appreciation of his leadership and contributions in all facets of antibiotic research. PrefaceThe idea for publishing these books on the mechanism of action and on the biosynthesis of antibiotics was born of frustration in our attempts to keep abreast of the literature. Gone were the years when we were able to keep a bibliography on antibiotics and feel confident that we could find everything that was being published on this subject. These fields of investigation were moving forward so rapidly and were encompassing so wide a range of specialized areas in microbiology and chemistry that it was almost impossible to keep abreast of developments. In our naivete and enthusiasm, however, we were unaware that we were toying with an idea that might enmesh us, that we were creating an entity with a life of its own, that we were letting loose a Golom who instead of being our servant would be our master.That we set up ideals for these books is obvious; they would be current guides to developments and information in the areas of mechanism of action and biosynthesis of antibiotics. For almost every subject, we wished to enlist the aid of an investigator who himself had played a part in determining the nature of the phenomena that were being discussed. One concept for the books was that they include only antibiotics for which a definitive, well-documented mechanism of action or biosynthetic pathway was known. Yet, such an approach would not entirely serve the purpose we projected, for it would not encompass all of the information available in these fields of antibiotic investigations and blind searches for the original literature would still have to be made. We therefore chose to include any and all antibiotics about which some pertinent information had been published. It was obvious even at the start that such a compilation, integration, and analysis of information could never be complete unless scientific investigations ceased at the moment the last manuscript was submitted-an end that was neither desirable nor possible. An addendum was therefore included at the end of the volume and left open for the addition of new information until the last pages of the regular articles had been printed.The original concept has also been further expanded as the authors of several papers found it advisable to elaborate on their subject. Some of the articles included discussions...

show abstract

Molecular mechanism of thymine-less death

Cited by 72 publications

References 5 publications

Accumulation of DNA strand breaks and methotrexate cytotoxicity.

Accumulation of DNA strand breaks and methotrexate cytotoxicity.

A genetical study of thymineless mutants ofE. coliK12

Antibiotics

Contact Info

Product

Resources

About