Repair Synthesis in Human Lymphocytes Provoked by Photolysis of Ethidium Azide

Cantrell, Charles E.; Yielding, K. Lemone

doi:10.1111/j.1751-1097.1977.tb06896.x

Cited by 20 publications

(4 citation statements)

References 8 publications

(3 reference statements)

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“…The ethidium photoaffinity probes have proved interesting in studies of mitochondrial mutagenesis (Hixon et al, 1975;Morita & Yielding, 1977Fukunaga & Yielding, 1979;Fukunaga et al, 1980), frame-shift mutagenesis (Yielding et al, 1976(Yielding et al, , 1979a; , and DNA repair (Cantrell & Yielding, 1977, 1981. Previously, the similarity of the in vivo binding sites of ethidium bromide, ethidium monoazide, and ethidium diazide was demonstrated by competition studies (Yielding et al, 1979b).…”

mentioning

confidence: 99%

Ethidium bromide and its photoreactive analogs: spectroscopic anlysis of deoxyribonucleic acid binding properties

Graves¹,

Watkins²,

Yielding³

1981

Biochemistry

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mentioning

confidence: 99%

Ethidium bromide and its photoreactive analogs: spectroscopic anlysis of deoxyribonucleic acid binding properties

Graves¹,

Watkins²,

Yielding³

1981

Biochemistry

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“…UV and ethidium azide photolysis on provoked repair synthesis (Cantrell and Yielding, 1977). Furthermore, comparable levels of ethidium bromide on pre-photolyzed ethidium azide did not inhibit UV repair.…”

Section: Previous Studies Did Not Show Additive Effects Ofmentioning

confidence: 82%

“…The covalent attachment of the drug moiety o n photolysis causes irreversible binding and thus provides a means of correlating a variety of biological processes with the kinetic and molecular details of drug binding. W e have employed photoaffinity labeling with an ethidium probe as a means of introducing repairable lesions into DNA of intact cells (Cantrell and Yielding, 1977). DNA repair synthesis in lymphocytes was used as a measure of the provoked repair and was found not t o be completely additive with UV damage either a t saturating or subsaturating doses of the azide.…”

Section: Introductionmentioning

confidence: 99%

Dna Excision Repair in Lymphocytes Following Photoaffinity Labeling With Ethidium Monoazide

Cantrell¹,

Yielding²

1980

Photochem & Photobiology

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Abstract— Irradiation of ethidium monoazide by fluorescent light promotes a chemical decomposition of the azide into a highly reactive nitrene intermediate. Covalent bonding of this electrophile to the DNA in the cell provokes repair of damage which can be monitored by incorporation of [3H]‐thymidine. Human lymphocytes were labeled with [14C]‐ethidium azide and then allowed to undergo DNA repair. Repair incorporation of [3H]‐thymidine showed saturation at 5 µM ethidium azide, but excision of the labeled drug failed to saturate at 20 µM, suggesting that excision and resynthesis are two separate events. Cells were also labeled with the photosensitive drug and/or exposed to UV radiation, and then allowed to undergo a period of DNA repair. The tritium incorporation for the combined insults was less than the sum of the two insults. Quinacrine, progesterone and chloroquine inhibited repair incorporation of [3H]‐thymidine, but had no effect on the excision of the drug from the DNA. After damage by ethidium azide, chromatin was isolated from lymphocytes which had been allowed to repair label with [3H]‐thymidine. Partial digestion of the chromatin with micrococcal nuclease released 80% of the tritium when approximately 40% of the DNA had been hydrolyzed by the enzyme.

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“…The specificity of such labeling is still a matter of interest, since histones do not display a significant intrinsic binding affinity for ethidium due presumably to their positive charges. 25 The occurrence of even a small amount of DNA-directed labeling of adjacent proteins could prove useful in studying the relationships of DNA and protein in intact chromatin. Substantial drug binding can occur to total nuclear proteins as reported previously for higher drug concentrati0ns.l' This could be due to a primary affinity for the drug rather than "noise" of the photolytic process for the DNA complex.…”

Section: Lack Of Dna Labeling Not Accounted For By Label On Proteinmentioning

confidence: 99%

Monoazido analog of ethidium as a chromatin probe: Binding to DNA

Coffman

Yielding

1984

Biopolymers

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SynopsisTwo photoaffinity analogs of ethidium, 8-azido-3-amin0, and 3-azido-8-amino-5-ethyl-6-phenylphenanthridinium chloride, have been used to probe the structure of mammalian chromatin and its interactions with the ethidium moiety. The monoazido analogs were established as suitable probes by comparing their interactions with chromatin and pure DNA prepared from chromatin to those of the parent ethidium bromide. Scatchard analysis of t h e binding data determined from spectrophotometric titrations showed that the analogs interacted with both nucleic acids in a manner similar to the parent compound. T h e effect of chromatin proteins on the interaction of the ethidium moiety with intact chromatin was investigated directly. By exposing the noncovalent complex to visible light, the monoazido analog was attached covalently in its interaction sites within chromatin, and the amount of drug bound covalently to DNA was determined for both protein-free DNA and chromatin. Using saturating concentrations of drug, DNA within intact chromatin was found to be associated with only half as much drug as DNA extracted from its protein prior to drug exposure. T h e distribution of drug bound within chromatin was determined following the attachment of the monoazido analog (by photoactivation) to chromatin that had undergone limited nuclease digestion. Several distinct populations isolated by size fractionation and quantitative measurements revealed that (1 ) both the core particles and the spacer-containing particles contained bound drug, reflecting high-affinity binding sites; and (2) chromatin particles containing 150 DNA base pairs (putatively nucleosome core structures) contained less total hound drug a t high drug concentrations than those particles having intact spacer DNA.

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Repair Synthesis in Human Lymphocytes Provoked by Photolysis of Ethidium Azide

Cited by 20 publications

References 8 publications

Ethidium bromide and its photoreactive analogs: spectroscopic anlysis of deoxyribonucleic acid binding properties

Ethidium bromide and its photoreactive analogs: spectroscopic anlysis of deoxyribonucleic acid binding properties

Dna Excision Repair in Lymphocytes Following Photoaffinity Labeling With Ethidium Monoazide

Monoazido analog of ethidium as a chromatin probe: Binding to DNA

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