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Title: Role of DNA polymerase. cap alpha. in chromosomal aberration production by ionizing radiation

Abstract

Aphidicolin is a tetracyclic diterpinoid fungal antibiotic which inhibits DNA synthesis in eukaryotic cells by interfering specifically with DNA polymerase ..cap alpha.., apparently by binding to and inactivating the DNA-polymerase ..cap alpha.. complex. We have shown that aphidicolin, like other inhibitors of DNA synthesis, both induces chromosomal aberrations in human peripheral lymphocytes, and, as a post-treatment, interacts synergistically with x rays to produce greatly enhanced aberration yields. The present experiments explore the effects of aphidicolin in human lymphocytes in the post-DNA-synthetic G/sub 2/ phase of the cell cycle. These experiments utilized labeling with tritiated thymidine to positively identify cells in the S phase at the time of treatment, and used serial colcemid collections and fixations to determine aberration yields over as much of the G/sub 2/ phase as feasible. Because DNA polymerase ..cap alpha.. is the only DNA synthetic or repair enzyme known to be affected by aphidicolin, we infer that this enzyme is directly involved in the repair of DNA lesions which can result in visible chromosomal aberrations. (DT)

Authors:
Publication Date:
Research Org.:
Brookhaven National Lab., Upton, NY (USA)
OSTI Identifier:
5478373
Report Number(s):
BNL-33857; CONF-8310231-2
ON: DE84002862
DOE Contract Number:
AC02-76CH00016
Resource Type:
Technical Report
Resource Relation:
Conference: International conference on hematopoietic cellular proliferation, Upton, NY, USA, 6 Oct 1983
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; 63 RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTANT EFFECTS ON LIVING ORGS. AND BIOL. MAT.; ANTIBIOTICS; BIOLOGICAL EFFECTS; DNA; BIOLOGICAL REPAIR; DNA POLYMERASES; BIOLOGICAL FUNCTIONS; INHIBITION; LYMPHOCYTES; BIOLOGICAL RADIATION EFFECTS; CHROMOSOMAL ABERRATIONS; CELL CYCLE; EXPERIMENTAL DATA; FUNGI; HEREDITARY DISEASES; MAN; MITOSIS; NERVOUS SYSTEM DISEASES; SYNERGISM; TRACER TECHNIQUES; TRITIUM COMPOUNDS; X RADIATION; ANIMAL CELLS; ANIMALS; ANTI-INFECTIVE AGENTS; BIOLOGICAL MATERIALS; BIOLOGICAL RECOVERY; BLOOD; BLOOD CELLS; BODY FLUIDS; CELL DIVISION; CONNECTIVE TISSUE CELLS; DATA; DISEASES; DRUGS; ELECTROMAGNETIC RADIATION; ENZYMES; FUNCTIONS; INFORMATION; IONIZING RADIATIONS; ISOTOPE APPLICATIONS; LABELLED COMPOUNDS; LEUKOCYTES; MAMMALS; MATERIALS; MUTATIONS; NUCLEIC ACIDS; NUCLEOTIDYLTRANSFERASES; NUMERICAL DATA; ORGANIC COMPOUNDS; PHOSPHORUS-GROUP TRANSFERASES; PLANTS; POLYMERASES; PRIMATES; RADIATION EFFECTS; RADIATIONS; RECOVERY; REPAIR; SOMATIC CELLS; TRANSFERASES; VERTEBRATES; 550301* - Cytology- Tracer Techniques; 560121 - Radiation Effects on Cells- External Source- (-1987)

Citation Formats

Bender, M.A. Role of DNA polymerase. cap alpha. in chromosomal aberration production by ionizing radiation. United States: N. p., 1983. Web. doi:10.2172/5478373.
Bender, M.A. Role of DNA polymerase. cap alpha. in chromosomal aberration production by ionizing radiation. United States. doi:10.2172/5478373.
Bender, M.A. Sat . "Role of DNA polymerase. cap alpha. in chromosomal aberration production by ionizing radiation". United States. doi:10.2172/5478373. https://www.osti.gov/servlets/purl/5478373.
@article{osti_5478373,
title = {Role of DNA polymerase. cap alpha. in chromosomal aberration production by ionizing radiation},
author = {Bender, M.A.},
abstractNote = {Aphidicolin is a tetracyclic diterpinoid fungal antibiotic which inhibits DNA synthesis in eukaryotic cells by interfering specifically with DNA polymerase ..cap alpha.., apparently by binding to and inactivating the DNA-polymerase ..cap alpha.. complex. We have shown that aphidicolin, like other inhibitors of DNA synthesis, both induces chromosomal aberrations in human peripheral lymphocytes, and, as a post-treatment, interacts synergistically with x rays to produce greatly enhanced aberration yields. The present experiments explore the effects of aphidicolin in human lymphocytes in the post-DNA-synthetic G/sub 2/ phase of the cell cycle. These experiments utilized labeling with tritiated thymidine to positively identify cells in the S phase at the time of treatment, and used serial colcemid collections and fixations to determine aberration yields over as much of the G/sub 2/ phase as feasible. Because DNA polymerase ..cap alpha.. is the only DNA synthetic or repair enzyme known to be affected by aphidicolin, we infer that this enzyme is directly involved in the repair of DNA lesions which can result in visible chromosomal aberrations. (DT)},
doi = {10.2172/5478373},
journal = {},
number = ,
volume = ,
place = {United States},
year = {Sat Jan 01 00:00:00 EST 1983},
month = {Sat Jan 01 00:00:00 EST 1983}
}

Technical Report:

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  • In the plasmodial slime mold Physarum polycephalum the nuclear divisions are synchronous. Synthesis of DNA occurs very shontly after the division and occupies a relatively brief period of the cell cycle. In contrast to the intermittent occurrence of DNA synthesis the formation of thymidlne triphosphate from non-phosphorylated precursors (orotic acid and thymidine) takes place throughout interphase. DNA-polymerase, the enzyme capable of polymerizing deoxynucleoside triphosphates in the presence of primer or template-DNA, has been demonstrated in extracts of molds harvested at any time of the cell cycle. Extracts made from molds irradiated during interphase with 100,000 r of x radiation showedmore » no decrease in polymerase activity as compared with extracts from unirradiated controls. It appears that in Physarum the DNA synthesis is not controlled by variations in the amount of DNA-polymerase and, furthermore that this enzyme is very insensitive to ionizing radiation. (auth)« less
  • Theoretical and experimental studies of free-radical yields in oriented DNA samples exposed to ionizing radiation with high linear energy transfer at 77 K are discussed. The dependence of radical yields on the orientation of DNA chains relative to the particle flux is being investigated to gain insight into the role of intramolecular energy and charge transfer processes in radical production and decay. Model calculations based on a thermal-spike approximation are presented and their limitations for predicting the orientation dependence of radical yields observed after neutron irradiation. Experiments on oriented DNA exposed to neutrons indicate that free-radical yields depend upon themore » orientation of DNA molecules relative to the recoil-proton flux. We developed a macroscopic model for this effect based on the assumption that vibrational energy migrates along DNA chains more rapidly than between DNA molecules in the irradiated sample, and approximated this difference by asymmetry in thermal-diffusion coefficients.« less
  • DNA polymerases delta and ..cap alpha.. were purified from CV-1 cells, and their sensitivities to the inhibitors aphidicolin, (p-n-butylphenyl) deoxyguanosine triphosphate (BuPdGTP), and monoclonal antibodies directed against DNA polymerase ..cap alpha.. were determined. The effects of these inhibitors on DNA replication, measured with (/sup 3/H)TTP, in permeabilized CV-1 cells were studied to investigate the potential roles of polymerases delta and ..cap alpha.. in DNA replication. Aphidicolin was shown to be a more potent inhibitor of DNA replication than of DNA polymerase ..cap alpha.. or delta activity. Inhibition of DNA replication by various concentrations of BuPdGTP was intermediate between inhibition ofmore » purified polymerase ..cap alpha.. or delta activity. Concentrations of BuPdGTP which totally abolished DNA polymerase ..cap alpha.. activity were much less effective in reducing DNA replication, as well as the activity of DNA polymerase delta. Monoclonal antibodies which specifically inhibited polymerase ..cap alpha.. activity reduced, but did not abolish, DNA replications in permeable cells. BuPdGTP, as well as anti-polymerase ..cap alpha.. antibodies, inhibited DNA replication in a nonlinear manner as a function of time. A concentration of BuPdGTP which abolished polymerase ..cap alpha.. activity reduced, but did not abolish, both the synthesis and maturation of nascent DNA fragments. This information suggests that polymerases ..cap alpha.. and delta are involved in both the synthesis and maturation of nascent DNA. This is the first report to present evidence suggesting that both polymerases ..cap alpha.. and delta play a significant role in mammalian DNA replication.« less
  • The levels of DNA polymerase ..cap alpha.. and ..beta.. activities were measured during the cell cycle using a total cell homogenate technique. The results indicate a decrease in the levels of both enzyme activities during G1 phase and a gradual increase as cells enter the S phase. The recovery of DNA polymerase activities was measured after heating for 10 min at 45.5/sup 0/C during the G1 phase or S phase. This treatment reduced the levels of DNA polymerase ..beta.. activity to 20-30% and DNA polymerase ..cap alpha.. activity to 40-50% of their control activity for both G1 and S phase.more » The activity of DNA polymerase ..beta.. recovered fully during 20-25 hr after heating for both G1 phase or S phase cells. There was no recovery of the activity of DNA polymerase ..cap alpha.. during this time. Survival was measured when cells were irradiated at various times after hyperthermia and for both G1 and S phase the interaction between heat and X rays disappeared by 20-25 hr after heating and the same increase was observed for the recovery of DNA polymerase ..beta.. activity. These results suggest that ..beta.. polymerase may be an important enzyme involved in repairing X-ray-induced damage that can result in cell lethality.« less