Uysal Özdemir, Özge2024-02-132024-02-132024-02-132022-11-3010.5075/epfl-UPSIM-307672https://infoscience.epfl.ch/handle/20.500.14299/203626All cells need to be able to sense their environment, and adapt their metabolism, growth and cell division appropriately. In this study, I examined the response of Schizosaccharomyces pombe to a change in the carbon source and how that affects cytokinesis. The focus of my work was the Septation Initiation Network (SIN), which is required for contractile actomyosin ring (CAR) assembly and septum formation. The work presented here stems from analysis of the genetic interaction network of etd1. Arguably, Etd1p is the least well-understood component of the SIN and its function in the SIN remains enigmatic. It is not essential at 36°C and at low temperatures, etd1-Δ dies with an elongated multinucleated phenotype. An extragenic suppressor of an etd1 mutant mapped to pdc101, the main pyruvate decarboxylase isoform in S. pombe. Since fission yeast grows mainly by fermentation in glucose (GLU) medium, Pdc101p plays an essential role the conversion of pyruvate to ethanol and the regeneration of NAD+ in the cytosol. We found that Pdc101p is essential at high temperatures in GLU medium. However, growth in glycerol (GLY) medium, a non-fermentable source, bypassed the requirement for Pdc101p, and surprisingly, Etd1p. GLY medium also rescued, but did not bypass the requirement for, several other SIN mutants. This unexpected result prompted us to investigate how carbon metabolism interacts with the SIN. We characterized wild-type cells in GLY medium and found that the cells are semi-wee, mimicking glucose-limitation. Glucose-sensing impacts mainly upon the cAMP/PKA pathway in S.pombe. Protein kinase A (Pka1p) is not essential for growth in GLU or GLY medium, though it is required for adjustment to a change of carbon source. Our data revealed that GLY medium decreased the intracellular cAMP level leading to inactivation of Pka1p; pka1-Δ cells in GLU medium largely phenocopy GLY grown wild-type cells. We found that GLY medium or deletion of pka1 increased the restrictive temperature of some of the SIN mutants, to the same extent. Our genetic analysis revealed that neither AMP-activated protein kinase nor the Greatwall-Endosulfine pathway is required for the rescue of the SIN mutants by GLY medium or pka1-Δ. Analysis of etd1-Δ and etd1-Δ pka1-Δ cells in GLU or GLY medium revealed that a significant proportion of etd1-Δ cells grown on GLU at 36°C (the permissive temperature) exhibit a cytokinesis defect. As expected, this proportion increased significantly at 29°C in GLU medium, but this was rescued if cells were grown in GLY medium or pka1 was deleted. Etd1p has an important role in the association of the SIN proteins with the SPBs, which is vital in the regulation of SIN signalling. Analysis of localisation of the SIN proteins in etd1-Δ confirmed that Etd1p is essential for Cdc7p localisation in late anaphase and the ring localisation of Mob1p in GLU medium at 29°C. We also found that a significant proportion of GLU-grown etd1-Δ cells lacked a GFP-Sid1p signal on the SPB during mitosis at both 36°C and 29°C. These defects were completely or partially rescued by growth in GLY medium or pka1 deletion, suggesting a role for Pka1p in controlling SPB association of SIN proteins. Interestingly, forced recruitment of Sid1p to the SPBs was also able to overcome the cytokinesis defects of etd1-Δ cells, and to rescue some SIN mutants. Our findings reveal novel aspects of SIN regulation and may challenge the current model of the SIN.enCellular BiologyCytologyGeneticsMetabolismdataset523e2e1a-5eaa-4872-bd4d-e4f0e0e94997