Abstract
Sporulation is a complex developmental program that fungi enter to ensure survival in unfavorable environmental conditions. Many fungal species are able to produce spores sexually through meiosis, which is beneficial since it introduces genetic variability into a population. The sexually reproducing ascomycete Saccharomyces cerevisiae has a well-defined sexual cycle, in which diploid cells can undergo meiosis and produce haploid spores in response to nutrient starvation. The diploid state is a requirement for meiosis and results from fusion of two haploid cells of the opposite mating type, which is regulated by the pheromone response pathway. Most ascomycetes have been reported to produce meiotic spores, however, a sexual cycle has not yet been identified in the filamentous fungus Ashbya gossypii. The main focus of my doctoral thesis has therefore been to understand the mechanisms behind sporulation in this fungus.
The lifecycle of A. gossypii starting with a haploid spore that matures into spore-containing mycelia can be completed without the need for a mating partner. Spores in A. gossypii are thought to be derived sexually like all other Saccharomycetaceae species, but the sexual cycle remains unidentified. In this thesis I provide a comprehensive functional analysis of genes important for sporulation in A. gossypii. Previous results, together with findings presented in this work show that the role of the pheromone response pathway in A. gossypii has been rewired to regulate sporulation negatively instead of controlling mating. In line with this, a mating partner might not be required since the multinucleate compartments could still enable nuclear fusion (karyogamy) and meiosis. The presence of karyogamy is supported by our results that deletion of the A. gossypii homologs Kar3 and Kar4, involved in karyogamy in S. cerevisiae, results in severely crippled sporulation.
This work also identifies the main regulators of sporulation in A. gossypiii namely IME1, IME2, IME4, KAR4 and NTD80. Using large scale RNA sequencing data of these non-sporulating deletion strains we were able to identify 67 down-regulated genes, most of which were up-regulated in an oversporulating mutant, namely ste12. Interestingly, transcription of the main regulator of meiosis in yeast, IME1, is regulated by a significantly smaller promoter in A. gossypii and is independent of the transcription factors Msn2 and Sok2. Furthermore, the role of Sok2 in A. gossypii has been rewired from a transcriptional repressor to an activator of sporulation. This is in contrast to S. cerevisiae, where Sok2 is a repressor of IME1 transcription while Msn2 and Msn4 function as activators.
A prerequisite for meiosis is meiotic recombination that allows cross-over between homologous chromosomes. In S. cerevisiae, this requires double strand break (DSB) formation and subsequent repair via the components Spo11 and Dmc1. The work in this thesis show that the A. gossypii Spo11 and Dmc1 homologs are not required for sporulation, thus suggesting that other proteins generate DSBs in this fungus.
In summary, this work has led to better understanding of the components regulating sporulation in A. gossypii and their hierarchical organization.
The lifecycle of A. gossypii starting with a haploid spore that matures into spore-containing mycelia can be completed without the need for a mating partner. Spores in A. gossypii are thought to be derived sexually like all other Saccharomycetaceae species, but the sexual cycle remains unidentified. In this thesis I provide a comprehensive functional analysis of genes important for sporulation in A. gossypii. Previous results, together with findings presented in this work show that the role of the pheromone response pathway in A. gossypii has been rewired to regulate sporulation negatively instead of controlling mating. In line with this, a mating partner might not be required since the multinucleate compartments could still enable nuclear fusion (karyogamy) and meiosis. The presence of karyogamy is supported by our results that deletion of the A. gossypii homologs Kar3 and Kar4, involved in karyogamy in S. cerevisiae, results in severely crippled sporulation.
This work also identifies the main regulators of sporulation in A. gossypiii namely IME1, IME2, IME4, KAR4 and NTD80. Using large scale RNA sequencing data of these non-sporulating deletion strains we were able to identify 67 down-regulated genes, most of which were up-regulated in an oversporulating mutant, namely ste12. Interestingly, transcription of the main regulator of meiosis in yeast, IME1, is regulated by a significantly smaller promoter in A. gossypii and is independent of the transcription factors Msn2 and Sok2. Furthermore, the role of Sok2 in A. gossypii has been rewired from a transcriptional repressor to an activator of sporulation. This is in contrast to S. cerevisiae, where Sok2 is a repressor of IME1 transcription while Msn2 and Msn4 function as activators.
A prerequisite for meiosis is meiotic recombination that allows cross-over between homologous chromosomes. In S. cerevisiae, this requires double strand break (DSB) formation and subsequent repair via the components Spo11 and Dmc1. The work in this thesis show that the A. gossypii Spo11 and Dmc1 homologs are not required for sporulation, thus suggesting that other proteins generate DSBs in this fungus.
In summary, this work has led to better understanding of the components regulating sporulation in A. gossypii and their hierarchical organization.
| Originalsprog | Engelsk |
|---|
| Forlag | Department of Biology, Faculty of Science, University of Copenhagen |
|---|---|
| Status | Udgivet - 2014 |