Increasing gene dosage provides a powerful means of probing gene function, as it tends to cause a gain-of-function effect due to increased gene activity. In the budding yeast, Saccharomyces cerevisiae, systematic gene overexpression studies have shown that in wild-type cells, overexpression of a small subset of genes results in an overt phenotype. However, examining the effects of gene overexpression in sensitized cells containing mutations in known genes is a powerful means for identifying functionally relevant genetic interactions. When a query mutant phenotype is rescued by gene overexpression, the genetic interaction is termed dosage suppression. I comprehensively investigated dosage suppression genetic interactions in yeast using three approaches. First, using one of two novel plasmid libraries cloned by two colleagues and myself, I systematically performed dosage suppression screens and identified over 130 novel dosage suppression genetic interactions for more than 25 essential yeast genes. The plasmid libraries, called the molecular barcoded yeast ORF (MoBY-ORF) 1.0 and 2.0, are designed to streamline dosage analysis by being compatible with high-throughput genomics technologies that can monitor plasmid representation, including barcode microarrays and next-generation sequencing methods. Second, I describe a detailed analysis of the novel dosage suppression interactions, as well as of literature-curated interactions, and show that the gene pairs exhibiting dosage suppression are often functionally related and can overlap with physical as well as negative genetic interactions. Third, I performed a systematic categorization of dosage suppression genetic interactions in yeast and show that the majority of the dosage suppression interactions can be assigned to one of four general mechanistic classifications. With this comprehensive analysis, I conclude that systematically identifying dosage suppression genetic interactions will allow for their integration into other genetic and physical interaction networks and should provide new insight into the global wiring diagram of the cell.
| Identifer | oai:union.ndltd.org:TORONTO/oai:tspace.library.utoronto.ca:1807/32169 |
| Date | 01 March 2012 |
| Creators | Magtanong, Leslie Joyce |
| Contributors | Boone, Charles |
| Source Sets | University of Toronto |
| Language | en_ca |
| Detected Language | English |
| Type | Thesis, Dataset |
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