Dataset Information

Combining Inhibitor Resistance-conferring Mutations in Cytochrome b Creates Conditional Synthetic Lethality in Saccharomyces cerevisiae.

ABSTRACT: The mitochondrial cytochrome bc(1) complex is an essential respiratory enzyme in oxygen-utilizing eukaryotic cells. Its core subunit, cytochrome b, contains two sites, center P and center N, that participate in the electron transfer activity of the bc(1) complex and that can be blocked by specific inhibitors. In yeast, there are various point mutations that confer inhibitor resistance at center P or center N. However, there are no yeast strains in which the bc(1) complex is resistant to both center P and center N inhibitors. We attempted to create such strains by crossing yeast strains with inhibitor-resistant mutations at center P with yeast strains with inhibitor-resistant mutations at center N. Characterization of yeast colonies emerging from the cross revealed that there were multiple colonies resistant against either inhibitor alone but that the mutational changes were ineffective when combined and when the yeast were grown in the presence of both inhibitors. Inhibitor titrations of bc(1) complex activities in mitochondrial membranes from the various yeast mutants showed that a mutation that confers resistance to an inhibitor at center P, when combined with a mutation that confers resistance to an inhibitor at center N, eliminates or markedly decreases the resistance conferred by the center N mutation. These results indicate that there is a pathway for structural communication between the two active sites of cytochrome b and open new possibilities for the utilization of center N as a potential drug target.

SUBMITTER: Ding MG

PROVIDER: S-EPMC2659206 | biostudies-literature | 2009 Mar

REPOSITORIES: biostudies-literature

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Combining Inhibitor Resistance-conferring Mutations in Cytochrome b Creates Conditional Synthetic Lethality in Saccharomyces cerevisiae.

Ding Martina G MG di Rago Jean-Paul JP Trumpower Bernard L BL

The Journal of biological chemistry 20090129 13

The mitochondrial cytochrome bc(1) complex is an essential respiratory enzyme in oxygen-utilizing eukaryotic cells. Its core subunit, cytochrome b, contains two sites, center P and center N, that participate in the electron transfer activity of the bc(1) complex and that can be blocked by specific inhibitors. In yeast, there are various point mutations that confer inhibitor resistance at center P or center N. However, there are no yeast strains in which the bc(1) complex is resistant to both cen ...[more]

PMID: 19179332

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Project description:BACKGROUND: Saccharomyces cerevisiae is the first eukaryotic organism for which a multi-compartment genome-scale metabolic model was constructed. Since then a sequence of improved metabolic reconstructions for yeast has been introduced. These metabolic models have been extensively used to elucidate the organizational principles of yeast metabolism and drive yeast strain engineering strategies for targeted overproductions. They have also served as a starting point and a benchmark for the reconstruction of genome-scale metabolic models for other eukaryotic organisms. In spite of the successive improvements in the details of the described metabolic processes, even the recent yeast model (i.e., iMM904) remains significantly less predictive than the latest E. coli model (i.e., iAF1260). This is manifested by its significantly lower specificity in predicting the outcome of grow/no grow experiments in comparison to the E. coli model. RESULTS: In this paper we make use of the automated GrowMatch procedure for restoring consistency with single gene deletion experiments in yeast and extend the procedure to make use of synthetic lethality data using the genome-scale model iMM904 as a basis. We identified and vetted using literature sources 120 distinct model modifications including various regulatory constraints for minimal and YP media. The incorporation of the suggested modifications led to a substantial increase in the fraction of correctly predicted lethal knockouts (i.e., specificity) from 38.84% (87 out of 224) to 53.57% (120 out of 224) for the minimal medium and from 24.73% (45 out of 182) to 40.11% (73 out of 182) for the YP medium. Synthetic lethality predictions improved from 12.03% (16 out of 133) to 23.31% (31 out of 133) for the minimal medium and from 6.96% (8 out of 115) to 13.04% (15 out of 115) for the YP medium. CONCLUSIONS: Overall, this study provides a roadmap for the computationally driven correction of multi-compartment genome-scale metabolic models and demonstrates the value of synthetic lethals as curation agents.

Dataset Information

Combining Inhibitor Resistance-conferring Mutations in Cytochrome b Creates Conditional Synthetic Lethality in Saccharomyces cerevisiae.

Publications

Combining Inhibitor Resistance-conferring Mutations in Cytochrome b Creates Conditional Synthetic Lethality in Saccharomyces cerevisiae.

Similar Datasets

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