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Genome Res. 2019 Jan 11. doi: 10.1101/gr.232330.117. [Epub ahead of print]

Transposon insertional mutagenesis in Saccharomyces uvarum reveals trans-acting effects influencing species-dependent essential genes.

Author information

1
Department of Genome Sciences, University of Washington, Seattle, Washington 98195, USA.
2
Molecular and Cellular Biology Program, University of Washington, Seattle, Washington 98195, USA.
3
Department of Biological Sciences, California State University, Turlock, California 95382, USA.
4
Energy Biosciences Institute, University of California Berkeley, Berkeley, California 94720, USA.
5
Buck Institute for Research on Aging, Novato, California 94945, USA.
6
Department of Plant and Microbial Biology, University of California Berkeley, Berkeley, California 94720, USA.
7
Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, Toronto, Ontario M5S 3E1, Canada.

Abstract

To understand how complex genetic networks perform and regulate diverse cellular processes, the function of each individual component must be defined. Comprehensive phenotypic studies of mutant alleles have been successful in model organisms in determining what processes depend on the normal function of a gene. These results are often ported to newly sequenced genomes by using sequence homology. However, sequence similarity does not always mean identical function or phenotype, suggesting that new methods are required to functionally annotate newly sequenced species. We have implemented comparative analysis by high-throughput experimental testing of gene dispensability in Saccharomyces uvarum, a sister species of Saccharomyces cerevisiae. We created haploid and heterozygous diploid Tn7 insertional mutagenesis libraries in S. uvarum to identify species-dependent essential genes, with the goal of detecting genes with divergent functions and/or different genetic interactions. Comprehensive gene dispensability comparisons with S. cerevisiae predicted diverged dispensability at 12% of conserved orthologs, and validation experiments confirmed 22 differentially essential genes. Despite their differences in essentiality, these genes were capable of cross-species complementation, demonstrating that trans-acting factors that are background-dependent contribute to differential gene essentiality. This study shows that direct experimental testing of gene disruption phenotypes across species can inform comparative genomic analyses and improve gene annotations. Our method can be widely applied in microorganisms to further our understanding of genome evolution.

PMID:
30635343
DOI:
10.1101/gr.232330.117

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