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Title: Identifying Genetic Interactions and Target Genes of Schizosaccharomyces pombe Transcription Factors by Functional Genomics
Author: Chatfield-Reed, Katherine
Advisor: Chua, Gordon
Keywords: Genetics;Microbiology;Biology--Molecular
Abstract: Transcriptional regulation of target genes is a critical component of cellular regulation. Changes in expression profiles characterize different cell types, stages of the cell cycle, reactions to the environment, and many diseases including cancer. Knowledge of transcriptional regulation is critical to understanding environmental adaption, cell fate, and targeted treatment of genetic diseases. Transcription factors do not act in isolation, and often have overlapping target genes or coordinated activity. As such, understanding transcription factors requires a global approach that can be achieved with high-throughput genomics. This study used Schizosaccharomyces pombe as a model organism to look at transcriptional regulation. Expression and chIP microarrays were used to look for the target genes of the calcineurin-responsive transcription factor Prz1. This work uncovered hundreds of putative target genes that were both positively and negatively regulated by Prz1. These genes illuminated an evolutionarily conserved function in the cell wall, and novel roles in flocculation and reproduction. The interplay between transcription factors was examined with a synthetic genetic array screen between the transcription factors. The double deletion mutants that were sicker suggest transcription factors that share target genes or regulate related processes. A full genome screen of Prz1 was used to look for possible genetic activators of Prz1. The pmr1+ calcium transporter gene was shown to negatively interact with prz1+, and increase Prz1 activity in the cell. While the alp31+ cofactor A gene also shared a negative genetic interaction with prz1+, it did not increase Prz1 activity. Finally, a synthetic dosage lethality screen was adapted for S. pombe and used to look for regulators of fourteen transcription factors. This screen found 195 sick interactions between the transcription factors and a miniarray of putative regulators. These interactions included two known upstream regulators of Yox1 and Scr1. It also showed interactions between Scr1 and two genes involved in protein degradation, the E3 ligase Ubr1 and the MYND domain protein SPBC31F10.10c, which are likely responsible for the ubiquitination of Scr1. These results introduce new insight into the S. pombe transcriptional-regulatory network, as well as providing a new methodology for examining genetic relationships in S. pombe in the future.
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