Chemotyping of yeast mutants using robotics

Rieger, Klaus; El-Alama, Mohamed; Stein, Georg; Bradshaw, Charles B.; Słonimski, Piotr P.; Maundrell, Kinsey

doi:10.1002/(sici)1097-0061(199907)15:10b<973::aid-yea402>3.0.co;2-l

Cited by 49 publications

(11 citation statements)

References 38 publications

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“…Accordingly, phenotypic effects can be uncovered by detailed comparisons of mutant and parental strains by using standardized PM assays. High-throughput assays based on 300 growth phenotypes have also been developed for Saccharomyces cerevisiae (65)(66)(67).…”

Section: Discussionmentioning

confidence: 99%

Phenotype MicroArray Analysis of Escherichia coli K-12 Mutants with Deletions of All Two-Component Systems

et al. 2003

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Two-component systems are the most common mechanism of transmembrane signal transduction in bacteria. A typical system consists of a histidine kinase and a partner response regulator. The histidine kinase senses an environmental signal, which it transmits to its partner response regulator via a series of autophosphorylation, phosphotransfer, and dephosphorylation reactions. Much work has been done on particular systems, including several systems with regulatory roles in cellular physiology, communication, development, and, in the case of bacterial pathogens, the expression of genes important for virulence. We used two methods to investigate two-component regulatory systems in Escherichia coli K-12. First, we systematically constructed mutants with deletions of all two-component systems by using a now-standard technique of gene disruption (K. A. Datsenko and B. L. Wanner, Proc. Natl. Acad. Sci. USA 97:6640-6645, 2000). We then analyzed these deletion mutants with a new technology called Phenotype MicroArrays, which permits assays of nearly 2,000 growth phenotypes simultaneously. In this study we tested 100 mutants, including mutants with individual deletions of all two-component systems and several related genes, including creBC-regulated genes (cbrA and cbrBC), phoBR-regulated genes (phoA, phoH, phnCDEFGHIJKLMNOP, psiE, and ugpBAECQ), csgD, luxS, and rpoS. The results of this battery of nearly 200,000 tests provided a wealth of new information concerning many of these systems. Of 37 different two-component mutants, 22 showed altered phenotypes. Many phenotypes were expected, and several new phenotypes were also revealed. The results are discussed in terms of the biological roles and other information concerning these systems, including DNA microarray data for a large number of the same mutants. Other mutational effects are also discussed.

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Section: Discussionmentioning

confidence: 99%

Phenotype MicroArray Analysis of Escherichia coli K-12 Mutants with Deletions of All Two-Component Systems

et al. 2003

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“…Further confirmation that the sensitivity was apparently due to another mutation was that the MATa deletant showed wildtype growth in the presence of oxidative stress (Figure 4). Approximately 80% of the phenotypes were confirmed by complementation with the cognate clones, a result that corresponds closely to those of Rieger et al (1999) for a variety of different chemotypes.…”

Section: Ynl099cmentioning

confidence: 61%

Phenotypic analysis of gene deletant strains for sensitivity to oxidative stress

Higgins¹,

Alic²,

Thorpe³

et al. 2002

Yeast

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Ascertaining the impact of inhibitors on the growth phenotype of yeast mutants can be useful in elucidating the function of genes within the cell. Microtitre plates and robotics have been used to screen over 600 deletions from EUROSCARF, constructed in an FY1679 strain background, for sensitivity to various oxidants. These included the inorganic hydroperoxide, H 2 O 2 , an organic peroxide (cumene hydroperoxide) and a lipid hydroperoxide (linoleic acid hydroperoxide). These produce within the cell several different reactive oxygen species that can cause damage to DNA, proteins and lipids. Approximately 14% of deletants displayed sensitivity to at least one of the oxidants and there was also a distribution of deletants that showed sensitivity to all or different combinations of the oxidants. Deletants included genes encoding proteins involved in stress responses, heavy metal homeostasis and putative cell wall proteins. Although global mechanisms have been identified that provide general stress responses, these results imply that there are also distinct mechanisms involved in the protection of the cell against specific damage caused by different oxidants. Further analysis of these genes may reveal unknown mechanisms protecting the cell against reactive oxygen species.

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“…Haploid strains with mutations in IES2, ISW1, RSC1, RSC2, SWR1, and YKU80 have been observed to display decreased growth in the presence of either methyl methonate sulfate (MMS), hydroxyurea (HU), or other DNA-damaging agents [19,[28][29][30][31]. The dosage dependence of the sensitivity was tested with the same set of haploid and diploid strains (Fig.…”

Section: Dosage Requirements For Dna Repairmentioning

confidence: 99%

The dosage of chromatin proteins affects transcriptional silencing and DNA repair in Saccharomyces cerevisiae

Benbow¹,

DuBois²

2008

FEBS Letters

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Alterations in protein composition or dosage within chromatin may trigger changes in processes such as gene expression and DNA repair. Through transposon mutagenesis and targeted gene deletions in haploids and diploids of Saccharomyces cerevisiae, we identified mutations that affect telomeric silencing in genes encoding telomere-associated Sir4p and Yku80p and chromatin remodeling ATPases Ies2p and Rsc1p. We found that sir4/SIR4 heterozygous diploids efficiently silence the mating type locus HMR but not telomeres, and diploids heterozygous for yku80 and ies2 mutations are inefficient at DNA repair. In contrast, strains heterozygous for most chromatin remodeling ATPase mutations retain wild-type silencing and DNA repair levels. Thus, in diploids, chromatin structures required for DNA repair and telomeric silencing are sensitive to dosage changes.

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Chemotyping of yeast mutants using robotics

Cited by 49 publications

References 38 publications

Phenotype MicroArray Analysis of Escherichia coli K-12 Mutants with Deletions of All Two-Component Systems

Phenotype MicroArray Analysis of Escherichia coli K-12 Mutants with Deletions of All Two-Component Systems

Phenotypic analysis of gene deletant strains for sensitivity to oxidative stress

The dosage of chromatin proteins affects transcriptional silencing and DNA repair in Saccharomyces cerevisiae

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