Abstract
When a replication fork collides with a DNA topoisomerase I (Top1) cleavage complex, the covalently bound enzyme must be removed from the DNA 3' end before recombination-dependent replication restart. Here we report that the tyrosyl-DNA phosphodiesterase Tdp1 and the structure-specific endonuclease Rad1-Rad10 function as primary alternative pathways of Top1 repair in Saccharomyces cerevisiae. Thus, tdp1 rad1 cells (including the catalytic point mutant rad1-D869A) not only are highly sensitive to the Top1 poison camptothecin but also exhibit a TOP1-dependent growth delay. Extensive genetic analysis revealed that both Tdp1 and Rad1-Rad10 repair proceed through recombination that equally depends on RAD52, RAD51, and RAD50. The Rad1-Rad10 pathway further particularly depends on RAD59 and SRS2 but is independent of other nucleotide excision repair genes. Although this pattern is consistent with Rad1-Rad10 removing Top1 in a manner similar to its removal of nonhomologous tails during gene conversion, these differ in that Top1 removal does not require Msh2-Msh3. Finally, we show that yeast lacking the Rad1-Rad10-related proteins Mus81-Mms4 display a unique pattern of camptothecin sensitivity and suggest a concerted model for the action of these endonucleases.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Camptothecin / pharmacology
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DNA Damage
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DNA Repair Enzymes
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DNA Repair* / drug effects
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DNA Repair* / genetics
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DNA Replication
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DNA Topoisomerases, Type I / metabolism*
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DNA, Fungal / metabolism
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DNA-Binding Proteins / metabolism
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Endonucleases / metabolism*
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Enzyme Inhibitors / pharmacology
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Flap Endonucleases
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Fungal Proteins / metabolism*
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Genes, Fungal
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Models, Biological
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Mutation
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Oligonucleotide Array Sequence Analysis
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Phosphoric Diester Hydrolases / metabolism*
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Recombination, Genetic
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Saccharomyces cerevisiae / drug effects
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Saccharomyces cerevisiae / genetics
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Saccharomyces cerevisiae / metabolism*
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Saccharomyces cerevisiae Proteins*
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Single-Strand Specific DNA and RNA Endonucleases
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Topoisomerase I Inhibitors
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Trans-Activators / metabolism
Substances
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DNA, Fungal
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DNA-Binding Proteins
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Enzyme Inhibitors
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Fungal Proteins
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Saccharomyces cerevisiae Proteins
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Topoisomerase I Inhibitors
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Trans-Activators
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Endonucleases
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Flap Endonucleases
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MUS81 protein, S cerevisiae
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RAD1 protein, S cerevisiae
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MMS4 protein, S cerevisiae
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RAD10 protein, S cerevisiae
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Single-Strand Specific DNA and RNA Endonucleases
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Phosphoric Diester Hydrolases
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tyrosyl-DNA phosphodiesterase
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DNA Topoisomerases, Type I
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DNA Repair Enzymes
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Camptothecin