Genome-wide screening for genes whose deletions confer sensitivity to mutagenic purine base analogs in yeast

Stepchenkova, Elena I.; Kozmin, Stanislav G.; Alenin, V. V.; Pavlov, Youri I.

doi:10.1186/1471-2156-6-31

Cited by 19 publications

(22 citation statements)

References 35 publications

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“…The YJL055W gene is known to confer resistance to 5-FOA when overexpressed, and further testing showed that it also confers resistance to 5-FU [19]. Conversely, a yjl055w mutant is sensitive to purine analogs [20]. It has therefore been proposed that YJL055W may function in detoxification of base analogs [19].…”

Section: Resultsmentioning

confidence: 99%

A Ham1p-Dependent Mechanism and Modulation of the Pyrimidine Biosynthetic Pathway Can Both Confer Resistance to 5-Fluorouracil in Yeast

Carlsson

Gustavsson

et al. 2013

PLoS ONE

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5-Fluorouracil (5-FU) is an anticancer drug and pyrimidine analogue. A problem in 5-FU therapy is acquired resistance to the drug. To find out more about the mechanisms of resistance, we screened a plasmid library in yeast for genes that confer 5-FU resistance when overexpressed. We cloned five genes: CPA1, CPA2, HMS1, HAM1 and YJL055W. CPA1 and CPA2 encode a carbamoyl phosphate synthase involved in arginine biosynthesis and HMS1 a helix-loop-helix transcription factor. Our results suggest that CPA1, CPA2, and HMS1 confer 5-FU resistance by stimulating pyrimidine biosynthesis. Thus, they are unable to confer 5-FU resistance in a ura2 mutant, and inhibit the uptake and incorporation into RNA of both uracil and 5-FU. In contrast, HAM1 and YJL055W confer 5-FU resistance in a ura2 mutant, and selectively inhibit incorporation into RNA of 5-FU but not uracil. HAM1 is the strongest resistance gene, but it partially depends on YJL055W for its function. This suggests that HAM1 and YJL055W function together in mediating resistance to 5-FU. Ham1p encodes an inosine triphosphate pyrophosphatase that has been implicated in resistance to purine analogues. Our results suggest that Ham1p could have a broader specificity that includes 5-FUTP and other pyrimidine analogoue triphosphates.

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Section: Resultsmentioning

confidence: 99%

A Ham1p-Dependent Mechanism and Modulation of the Pyrimidine Biosynthetic Pathway Can Both Confer Resistance to 5-Fluorouracil in Yeast

Carlsson

Gustavsson

et al. 2013

PLoS ONE

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“…The results of numerous global screens of sensitivity to various genotoxic stress are also available [9], [42]. There is only modest overlap of our gene list with any of the published studies, but they are also quite dissimilar (see Supplementary Table S2).…”

Section: Discussionmentioning

confidence: 98%

Dipoid-Specific Genome Stability Genes of S. cerevisiae: Genomic Screen Reveals Haploidization as an Escape from Persisting DNA Rearrangement Stress

2011

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Maintaining a stable genome is one of the most important tasks of every living cell and the mechanisms ensuring it are similar in all of them. The events leading to changes in DNA sequence (mutations) in diploid cells occur one to two orders of magnitude more frequently than in haploid cells. The majority of those events lead to loss of heterozygosity at the mutagenesis marker, thus diploid-specific genome stability mechanisms can be anticipated. In a new global screen for spontaneous loss of function at heterozygous forward mutagenesis marker locus, employing three different mutagenesis markers, we selected genes whose deletion causes genetic instability in diploid Saccharomyces cerevisiae cells. We have found numerous genes connected with DNA replication and repair, remodeling of chromatin, cell cycle control, stress response, and in particular the structural maintenance of chromosome complexes. We have also identified 59 uncharacterized or dubious ORFs, which show the genome instability phenotype when deleted. For one of the strongest mutators revealed in our screen, ctf18Δ/ctf18Δ the genome instability manifests as a tendency to lose the whole set of chromosomes. We postulate that this phenomenon might diminish the devastating effects of DNA rearrangements, thereby increasing the cell's chances of surviving stressful conditions. We believe that numerous new genes implicated in genome maintenance, together with newly discovered phenomenon of ploidy reduction, will help revealing novel molecular processes involved in the genome stability of diploid cells. They also provide the clues in the quest for new therapeutic targets to cure human genome instability-related diseases.

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“…It is possible that the DIAL probe may be a substrate for some minor activity, particularly at long reaction times as seen here. Several other genes, in addition to HAM1 , are involved in protection from HAP in yeast ( 62 ). Future characterization of background activity in extracts of yeast double mutants with deletion of ITPA and these genes or genes encoding other phosphatases will shed light on additional enzymes that participate in cleansing the nucleotide pool of deamination damage in eukaryotes.…”

Section: Discussionmentioning

confidence: 99%

Measuring deaminated nucleotide surveillance enzyme ITPA activity with an ATP-releasing nucleotide chimera

Stepchenkova

Cui

et al. 2017

Nucleic Acids Research

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Nucleotide quality surveillance enzymes play important roles in human health, by detecting damaged molecules in the nucleotide pool and deactivating them before they are incorporated into chromosomal DNA or adversely affect metabolism. In particular, deamination of adenine moiety in (deoxy)nucleoside triphosphates, resulting in formation of (d)ITP, can be deleterious, leading to DNA damage, mutagenesis and other harmful cellular effects. The 21.5 kDa human enzyme that mitigates this damage by conversion of (d)ITP to monophosphate, ITPA, has been proposed as a possible therapeutic and diagnostic target for multiple diseases. Measuring the activity of this enzyme is useful both in basic research and in clinical applications involving this pathway, but current methods are nonselective and are not applicable to measurement of the enzyme from cells or tissues. Here, we describe the design and synthesis of an ITPA-specific chimeric dinucleotide (DIAL) that replaces the pyrophosphate leaving group of the native substrate with adenosine triphosphate, enabling sensitive detection via luciferase luminescence signaling. The probe is shown to function sensitively and selectively to quantify enzyme activity in vitro, and can be used to measure the activity of ITPA in bacterial, yeast and human cell lysates.

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Genome-wide screening for genes whose deletions confer sensitivity to mutagenic purine base analogs in yeast

Cited by 19 publications

References 35 publications

A Ham1p-Dependent Mechanism and Modulation of the Pyrimidine Biosynthetic Pathway Can Both Confer Resistance to 5-Fluorouracil in Yeast

A Ham1p-Dependent Mechanism and Modulation of the Pyrimidine Biosynthetic Pathway Can Both Confer Resistance to 5-Fluorouracil in Yeast

Dipoid-Specific Genome Stability Genes of S. cerevisiae: Genomic Screen Reveals Haploidization as an Escape from Persisting DNA Rearrangement Stress

Measuring deaminated nucleotide surveillance enzyme ITPA activity with an ATP-releasing nucleotide chimera

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