Elevated levels of sphingolipid MIPC in the plasma membrane disrupt the coordination of cell growth with cell wall formation in fission yeast

Coupling cell wall expansion with cell growth is a universal challenge faced by walled organisms. Mutations in Schizosaccharomyces pombe css1, which encodes a PM inositol phosphosphingolipid phospholipase C, prevent cell wall expansion but not synthesis of cell wall material. To probe how Css1 modulates cell wall formation we used classical and chemical genetics coupled with quantitative mass spectrometry. We found that elevated levels of the sphingolipid biosynthetic pathway's final product, mannosylinositol phosphorylceramide (MIPC), specifically correlated with the css1-3 phenotype. We also found that an apparent indicator of sphingolipids and a sterol biosensor accumulated at the cytosolic face of the PM at cell tips and the division site of css1-3 cells and, in accord, the PM in css1-3 was less dynamic than in wildtype cells. Interestingly, disrupting the protein glycosylation machinery recapitulated the css1-3 phenotype and led us to investigate Ghs2, a glycosylated PM protein predicted to modify cell wall material. Disrupting Ghs2 function led to aberrant cell wall material accumulation suggesting Ghs2 is dysfunctional in css1-3. We conclude that preventing an excess of MIPC in the S. pombe PM is critical to the function of key PM-localized proteins necessary for coupling growth with cell wall formation. Many organisms have a cell wall that provides structural support and serves as protection against environmental stresses. At the same time, cell walls are dynamic, needing to be repaired and elaborated on as cells grow and divide. A shared challenge among cell-walled organisms is therefore maintaining the integrity of the cell wall while it is being remodeled. The yeast Schizosaccharomyces pombe has a well understood pattern of new cell wall deposition at growing tips during interphase and along the septum at division. We took advantage of this knowledge to study a poorly understood step in cell wall assembly defined by a mutation in the css1 gene that encodes a sphingomyelinase-like enzyme. In these cells, a step following cell wall precursor synthesis fails and therefore, cells stop elongating. We determined that changes in plasma membrane lipid composition and dynamics accompany this defect and identify putative enzymes that act on cell wall precursors after their synthesis to allow cell growth.