Schizosaccharomyces pombe Phosphatidylinositol 4-kinase, Pik1p, in cell cycle control

Abstract

Pik1p, one of three phosphatidylinositol 4-kinases in the fission yeast, Schizosaccharomyces pombe, was found previously to interact with Cdc4p, a myosin essential light chain that is required for cytokinesis. The involvement of pik1 in cell cycle control was investigated. A fluorescently tagged Pik1p fusion protein was associated with Golgi throughout the cycle, and was found at the medial division plane of the cell during late cytokinesis. This latter distribution has not been reported previously. Gene deletion in diploid cells and tetrad analysis revealed that pik1 is essential for cell viability and is required for spore germination. The terminal phenotype of a temperature-sensitive, loss-of-function allele (pik1-td) indicated that pik1 is involved in cytokinesis: particularly for suppression of secondary septum material deposition, for suppression of initiation of supernumerary septa, and for cell separation. Contractile ring formation was normal in pik1-td cells at the restrictive temperature although the pattern of F-actin patches was disrupted. The F-actin patches were dispersed throughout the cytoplasm. Accumulation of extra inner membranous or vesicle-like structures was observed in these cells. The S. pombe nmt1 promoter and attenuated versions of it were found to be useful for complementation studies in S. cerevisiae. Heterologous expression of S. pombe pik1 complemented the essential functions of a temperature-sensitive allele (pik1-101) of its orthologue in Saccharomyces cerevisiae that were lost at the restrictive temperature. A residue required for S. pombe Pik1p lipid kinase activity, D709, was also required for this complementation. A residue, R838, which is required for interactions between Pik1p and Cdc4p was not required for this complementation. The timing and localization of Pik1p to the division plane of the cell late in cytokinesis combined with analysis of the terminal phenotype of a loss-of-function allele, indicate that Pik1p and/or its derived phosphoinositides are required for regulation of septation and cell separation. Pik1p may be involved in the transport, possibly via vesicular transport, of enzymes required for hydrolysis of the primary septum. It may be involved in signaling pathways that lead to the initiation of septation and to the cessation of the deposition of secondary septum material.