CHARACTERIZATION OF HIGH AFFINITY ACTINOMYCIN D BINDING TO EUKARYOTIC DNA

Persistent Link:
http://hdl.handle.net/10150/282405
Title:
CHARACTERIZATION OF HIGH AFFINITY ACTINOMYCIN D BINDING TO EUKARYOTIC DNA
Author:
Khan, Manzoor Mahmood
Issue Date:
1980
Publisher:
The University of Arizona.
Rights:
Copyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.
Abstract:
Actinomycin D in low concentrations was suggested to inhibit ribosomal RNA (rRNA) transcription via an extranucleolar mechanism. Actinomycin D was proposed to inhibit unique messenger RNAs (mRNAs) coding for proteins needed for the maintenance of rRNA transcription. According to this hypothesis actinomycin D would bind to specific nonribosomal DNA with high affinity. This hypothesis was investigated by isolating high molecular weight rat liver DNA, digesting it with restriction endonuclease EcoRI, adding [³H] actinomycin D in low concentration, performing RPC-5 chromatography to separate the restriction fragments and subsequent hybridization to rRNA. It was observed that actinomycin D bound to nonribosomal DNA with high affinity. The same experiment was performed with nucleolar DNA. High affinity actinomycin D binding was not observed in nucleolar DNA. Discrete high affinity binding DNA for actinomycin in rat liver DNA was also observed when another restriction endonuclease BamHI was used to cleave rat liver DNA. However, with rat liver DNA digested with restriction endonuclease HindIII, such a high affinity actinomycin D binding DNA was not observed. Actinomycin D was also demonstrated to bind to discrete site(s) in at least four more eukaryotic species (salmon, calf, herring and human) after DNA from these species were digested by EcoRI, labeled actinomycin D added, and RPC-5 chromatography performed. Labeled actinomycin D bound to its high affinity binding DNA was displaced by unlabeled actinomycin D in a concentration range of biological significance. However, six other antitumor agents, (doxorubicin, aclacinomycin, carminomycin, marcellomycin, musettamycin and pyrromycin) which also intercalate into DNA, did not significantly displace labeled actinomycin D from its high affinity binding DNA. Since this high affinity actinomycin D binding DNA is hypothesized to be involved in the inhibition of rRNA transcription, the actinomycin D binding DNA could have a role in the regulation of rRNA transcription. To date this is the first time that a probable regulatory DNA has been characterized by selective drug binding.
Type:
text; Dissertation-Reproduction (electronic)
Keywords:
Actinomycin.; Antibiotics -- Testing.; DNA.
Degree Name:
Ph.D.
Degree Level:
doctoral
Degree Program:
Graduate College; Pharmacology
Degree Grantor:
University of Arizona
Advisor:
Lindell, Thomas

Full metadata record

DC FieldValue Language
dc.language.isoen_USen_US
dc.titleCHARACTERIZATION OF HIGH AFFINITY ACTINOMYCIN D BINDING TO EUKARYOTIC DNAen_US
dc.creatorKhan, Manzoor Mahmooden_US
dc.contributor.authorKhan, Manzoor Mahmooden_US
dc.date.issued1980en_US
dc.publisherThe University of Arizona.en_US
dc.rightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.en_US
dc.description.abstractActinomycin D in low concentrations was suggested to inhibit ribosomal RNA (rRNA) transcription via an extranucleolar mechanism. Actinomycin D was proposed to inhibit unique messenger RNAs (mRNAs) coding for proteins needed for the maintenance of rRNA transcription. According to this hypothesis actinomycin D would bind to specific nonribosomal DNA with high affinity. This hypothesis was investigated by isolating high molecular weight rat liver DNA, digesting it with restriction endonuclease EcoRI, adding [³H] actinomycin D in low concentration, performing RPC-5 chromatography to separate the restriction fragments and subsequent hybridization to rRNA. It was observed that actinomycin D bound to nonribosomal DNA with high affinity. The same experiment was performed with nucleolar DNA. High affinity actinomycin D binding was not observed in nucleolar DNA. Discrete high affinity binding DNA for actinomycin in rat liver DNA was also observed when another restriction endonuclease BamHI was used to cleave rat liver DNA. However, with rat liver DNA digested with restriction endonuclease HindIII, such a high affinity actinomycin D binding DNA was not observed. Actinomycin D was also demonstrated to bind to discrete site(s) in at least four more eukaryotic species (salmon, calf, herring and human) after DNA from these species were digested by EcoRI, labeled actinomycin D added, and RPC-5 chromatography performed. Labeled actinomycin D bound to its high affinity binding DNA was displaced by unlabeled actinomycin D in a concentration range of biological significance. However, six other antitumor agents, (doxorubicin, aclacinomycin, carminomycin, marcellomycin, musettamycin and pyrromycin) which also intercalate into DNA, did not significantly displace labeled actinomycin D from its high affinity binding DNA. Since this high affinity actinomycin D binding DNA is hypothesized to be involved in the inhibition of rRNA transcription, the actinomycin D binding DNA could have a role in the regulation of rRNA transcription. To date this is the first time that a probable regulatory DNA has been characterized by selective drug binding.en_US
dc.typetexten_US
dc.typeDissertation-Reproduction (electronic)en_US
dc.subjectActinomycin.en_US
dc.subjectAntibiotics -- Testing.en_US
dc.subjectDNA.en_US
thesis.degree.namePh.D.en_US
thesis.degree.leveldoctoralen_US
thesis.degree.disciplineGraduate Collegeen_US
thesis.degree.disciplinePharmacologyen_US
thesis.degree.grantorUniversity of Arizonaen_US
dc.contributor.advisorLindell, Thomasen_US
dc.identifier.proquest8022829en_US
dc.identifier.oclc7546773en_US
dc.identifier.bibrecord.b18069642en_US
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