| The genetic interactions usually exist in a group of genes with functional relationships,and the relationships reflected in the the phenotypic changes of cells or organisms. A genetic interaction between two genes indicates that the phenotype of a double mutant differs from what is expected from each individual mutant. There is a positive correlation between the phenotypic difference and the intensity of genetic interaction. Screening of genetic interaction is an important method for understanding correlations between different genes, but also provide the clues for the deep insight of biological process as well as studying the potential role of a specific gene.SAGA complex is a highly conserved chromatin remodeling complex in eukaryotes, including 5 different functional modules formed by 21 subunit such as acetylation module and Dub module. It plays role in many important cellular processes, including transcriptional activation, histone modification and mRNA exportation. Most of the earlier studies focused on exploring the biological functions of SAGA complex as a whole and its different modules, on the contrary, less attention were paid on specific SAGA subunit. Sgf73 is a SAGA subunit, which highly conserved from yeast to human and constituted with Ubp8, Sgf11, Susl as the Dub module of SAGA complex. Due to the late funding of Sgf73, how it affects vital biological process in fission yeast is yet to be elucidated. In this study, we will excavate all the non-essential genes having genetic interactions with sgf73+ through screening method in fission yeast, and then investigate the molecular mechanism and potential biological function of sgf73+ in vivo.We chose the sgf73+ as a query gene to perform a global genetic interaction analysis through synthetic genetic array method with fission yeast non-essential gene deletion library consists of 3256 single mutants. The data suggest that 206 genes have genetic interactions with sgf73+, including 164 negative genetic interactions and 42 positive genetic interactions. We found that 74 genes having genetic interactions with sgf73+for the first time. GO term analysis illustrates that these genes are enriched in several important biological processes, including chromatin modification, DNA damage repair, cellular response to stress, transcription, cell cycle, signal transduction, transport and metabolism.This result prompts that sgf73+may have effect on these biological processes.Histone modification is a reversible post-translational process closely related to transcription and DNA damage repair. sgf73+ have genetic interactions with 19 chromatin modification related genes.The histone modification results showed that loss of sgf73+ led to the decreased levels of histone acetylation at H3K9 and H4K16, the decreased percentage at H3K9 was 20% and at H4K16 30%. The deletion of sgf73+ could also cause an increased level of histone H3K4 methylation. Compared with the loss of H3K4 methylation in ash2A strain, the methylation level in sgf73Δ ash2Δ double mutants strain recovered to the same level of wild type. This is the first time to identify that loss of sgf73+lead to the decreased levels of histone acetylation at H3K9 and H4K16. the increased level of histone H3K4 methylation in fission yeast.Furthermore,sgf73+ also showed genetic relationships with genes involved in DNA damage repair and stress responses. The spot assay results indicated that the sgf73Δ cells were sensitive to DNA damage agents HU,CPT,and H2O2 to some extent. In addition to. sgf73+ functioned in the same genetic pathway with rad51+, a gene related to DNA recombination repair process, suggesting sgf73+ may play a novel role in DNA damage repair.The TBZ sensitivity test on sgf73+and microtubule related genes klp6+, tipl+, mal3+ showed that sgf73+ was sensitive to TBZ independent of the SAGA complex, and stay in the same way with klp6+. Fluorescent microscopic analysis suggested that microtubule became shorter in sgf73Δ, and overexpressing Klp6 could significantly recover the change, suggesting sgf73+ acts upstream of klp6+ in regulating dynamic stability of microtubules.Taken together, we identified 206 genes having genetic interactions with sgf73+, and further studies indicated that as a multifunctional subunit of SAGA complex, Sgf73 played a key role in regulating histone modification, DNA damage repair and dynamic stability of microtubules. |
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