Abstract/Details

Mutant DNA topoisomerase I-induced lethality in Saccharomyces cerevisiae


1996 1996

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Abstract (summary)

Eukaryotic DNA topoisomerase I catalyzes the relaxation of supercoiled DNA through a concerted mechanism of DNA strand breakage and religation. The antineoplastic activity of camptothecin results from the specific and reversible stabilization of a covalent enzyme-DNA intermediate. Mutations in two highly conserved regions of Saccharomyces cerevisiae DNA topoisomerase I produced a similar mechanism of cell killing, in the absence of camptothecin. In Top1T722Ap, substitution of threonine-722 with alanine reduced enzyme specific activity about 3-fold, yet dramatically increased the stability of the covalent enzyme-DNA complex. The specific activity of Top1R517Gp was reduced about 1,000-fold; however, the cytotoxic action of this mutant protein could also be attributed to enhanced DNA cleavage. Overexpression of these lethal top1 mutants resulted in the accumulation of large-budded cells arrested in G2 of the cell cycle, while sub-lethal levels of top1T722A expression increased rDNA recombination. Mutation of the active site tyrosine completely abrogated the cytotoxicity and in vivo activity of the mutant enzymes, while deletion of the RAD9 cell cycle checkpoint enhanced top1 mutant-induced cell killing. The formation of these lethal, drug-independent DNA lesions are discussed in terms of the biological functions of eukaryotic DNA topoisomerase I.

Indexing (details)


Subject
Genetics;
Molecular biology
Classification
0369: Genetics
0307: Molecular biology
Identifier / keyword
Biological sciences; camptothecin
Title
Mutant DNA topoisomerase I-induced lethality in Saccharomyces cerevisiae
Author
Megonigal, Maureen D.
Number of pages
198
Publication year
1996
Degree date
1996
School code
0272
Source
DAI-B 59/04, Dissertation Abstracts International
Place of publication
Ann Arbor
Country of publication
United States
ISBN
0591817144, 9780591817140
Advisor
Bjornsti, Mary-Ann; Prockop, Darwin J.
University/institution
Thomas Jefferson University
University location
United States -- Pennsylvania
Degree
Ph.D.
Source type
Dissertations & Theses
Language
English
Document type
Dissertation/Thesis
Dissertation/thesis number
9829082
ProQuest document ID
304315292
Copyright
Database copyright ProQuest LLC; ProQuest does not claim copyright in the individual underlying works.
Document URL
http://search.proquest.com/docview/304315292
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