The involvement of inositol lipids and phosphates in signalling in the fission yeast Schizosaccharomyces pombe

Abstract

Inositol lipids and phosphates play vital roles in the control of cell function through their participation in signalling pathways downstream of receptor-regulated enzymes such as phospholipase C and phosphatidylinositol 3-kinase, which generate established (inositol 1,4,5-trisphosphate and diacylglycerol) or putative (phosphatidylinositol 3,4,S-trisphosphate) intracellular signalling molecules respectively (1 -3). Further progress in our understanding of the functions of these second messengers in complex cellular processes may come by studying signalling pathways in organisms with easily controlled genetics.The fission yeast, Schizosaccharomycespombe, in which mating is controlled by the reciprocal release of the pheromones M-factor and P-factor (4), may provide a genetically tractable organism in which to study these events. The pheromone response pathway appears to include certain features similar to well characterised signal transduction events in mammalian systems. The receptors for both pheromones (encoded by themam2 and map3 genes) have seven transmembrane spanning domains and are members of the receptor superfamily whose prototype is mammalian rhodopsin. Like other receptors of this family, the pheromone receptors are coupled to a G protein (encoded bygpal). The Gcr sub-unit is responsible for transducing the pheromone signal (5). S. Pombe also contain homologues of the mammalian MAP kinase cascade (byr2, byrl, ste8) and of ras (Rasl) and these proteins are also believed to be components of the pheromone signalling pathway, although their precise functions have yet to be defined (4) The direct target for the Gpal protein is unknown and has been the focus of our investigations. Preliminary experiments analysing the effect of pheromone stimulation of S. pombe cells have implicated the involvement of certain inositol lipids and phosphates in the signalling pathway. The most consistent response to pheromone was a triphasic accumulation of diacylglycerol (DAG), shown in Figure 1; this was measured by a mass assay involving radioenzymatic conversion of DAG to phosphatidic acid (PtdOH) by E.coli DAG kinase (6). In addition, increases in compounds having the chromatographic characteristics of glycerophosphoinositol and InsP, although not consistently in the levels of Ins( 1,4,5)P3, were observed upon pheromone stimulation in [3H]inositol-labelled S. pombe cells. Such observations implicate the activation of specific phospholipases in response to pheromone addition. Extracts from S. pombe contain a number of the inositol phosphates commonly found in mammalian cells, alongside some of the enzymes which metabolise them. Exogenously added [3H]Ins( 1,4,5)P3, [3H]Ins(1,4)P2 and [3H]Ins1P can be metabolised by activities which appear to share some of the characteristics of their mammalian counterparts (7). For example [3H]Ins( 1,4,5)P3 was metabolised by a 5-phosphatase, a 3-kinase and also by a minor pathway involving a putative 6-kinase. [3H]Ins( I,3,4)P3 appears to be dephosphorylated to [3H]Ins( 1,4)P2 and [3H]Ins(...

Inositol hexakisphosphate in Schizosaccharomyces pombe: synthesis from Ins(1,4,5)P3 and osmotic regulation

“…Neurospora crassa, Saccharomyces cere isiae, Schizosaccharomyces pombe), in which inositol utilization has been analysed extensively [16][17][18][19][20][21][22][23][24][25][26]. The stress-regulated PtdIns(3,5)P # pathway was described in [10], and yeast inositol lipids and\or phosphates may have roles in cell cycle control and responses to nutrient and osmotic stresses and mating pheromones [22][23][24][25][26]. It is still not clear whether yeast Ins(1,4,5)P $ is made by the one phosphoinositidase C present in these organisms (homologous to mammalian phosphoinositidase C-δ [16][17][18][19]) or by some other route, and whether it has a regulatory role.…”

Isolation of the plc1 gene from the fission yeast Schizosaccharomyces pombe

“…The signalling pathway is reasonably well understood but there is little information available concerning events between the G protein and the kinase cascade. Preliminary experiments have implicated the activation of specific phospholipases in response to pheromone addition [2] and we have used a PCR-based approach to identify genes that encode these enzymes ( Figure 1). We now report the identification of p l c l , the first S.pombe gene with the capacity to encode a phopsholipase C (PLC).…”