Characterization of the meiotic regulation of Superoxide dismutase 1 in Saccharomyces cerevisiae

Meiosis is the conserved cell division process used by sexually reproducing organisms to produce gametes, which are the cells that give rise to the next generation. In humans, meiosis generates eggs and sperm, and in the model budding yeast Saccharomyces cerevisiae, meiosis generates spores. Despite considerable evolutionary divergence, the general mechanics of meiotic chromosome division are largely conserved from yeast to humans. This conservation has made yeast an invaluable tool for understanding meiosis. In addition to important discoveries related to chromosome division, yeast have shaped our knowledge of other meiotic processes, including gene regulatory changes, organelle dynamics and inheritance, protein quality control, and cellular rejuvenation./nChanges in expression impact virtually every gene in the yeast genome during meiosis and spore development. In addition to increases and decreases in the transcription of classically defined genes and mRNAs, meiotic gene expression also involves the transcription of extended transcripts called Long Undecoded Transcript Isoforms (LUTIs), which have 5' extended sequences relative to their canonical counterparts. LUTIs are repressive, acting to transcriptionally interfere with canonical mRNA expression and translationally block protein synthesis from their main ORF via competitive translation of upstream open reading frames (uORFs). LUTIs are also pervasive and used to drive protein levels for approximately 8% of all yeast genes during meiosis. The majority of LUTI mRNAs have yet to be characterized in detail, and many of the proteins impacted by their expression are reported to exhibit stable expression in mitosis./nSuperoxide dismutase 1 (Sod1) is a highly abundant and primarily cytosolic antioxidant that detoxifies superoxide radicals (O2•–), a type of reactive oxygen species (ROS), from cells. We became interested in studying Sod1 after analysis of mRNA-sequencing and ribosome profiling data that suggested a LUTI mRNA is expressed from the SOD1 locus during meiosis. During mitotic growth, yeast express a canonical SOD1 mRNA (SOD1canon.) of approximately 600 nucleotides. During the meiotic divisions, cells initiate transcription from an extremely distal start site, resulting in the production of a putative LUTI mRNA that is nearly four times the length of SOD1canon.. We confirmed that this extended isoform (SOD1LUTI) is in fact produced as a continuous, ~2.2 kb transcript, and that LUTI expression functions to repress canonical mRNA levels. Although naturally produced during meiosis, SOD1LUTI is also made by mitotic cells experiencing ER stress, raising the possibility that the meiotic unfolded protein response (UPRER) is responsible for turning on the LUTI./nIn addition to highly dynamic changes in the expression of SOD1canon. and SOD1LUTI, we found that Sod1 protein levels and localization change during meiosis. During the transition from nutrient-rich media to starvation conditions, Sod1 levels decrease considerably and discrete foci of Sod1 appear. As cells go through meiosis, the brightness and number of these foci decreases, suggesting their degradation. The formation of these foci is independent of entry into meiosis, but their disappearance from cells relies on meiotic progression. During the meiotic divisions, due to the combination of LUTI expression and the degradation of preexisting protein, Sod1 levels reach their lowest levels. Once the chromosome divisions have completed, cells begin to express SOD1canon. once more, and new Sod1 protein is produced. Altogether, these data support a model in which old Sod1 protein is systematically cleared from cells to facilitate the generation of new protein. We propose that the regulation of this important antioxidant outlined in this dissertation may play a role the lifespan resetting that takes place during gametogenesis./nMany important questions remain unanswered regarding the meiotic regulation of Sod1. What is the significance of this transient dip in expression during meiosis? What is the nature of the relationship between SOD1LUTI and the UPRER? Are the foci of Sod1 we observe true protein aggregates? Answering these and other questions will improve our understanding of the biology of this important antioxidant.

Advisor: Brar, Gloria A