Molecular cloning of the plc1+ gene of Schizosaccharomyces pombe, which encodes a putative phosphoinositide-specific phospholipase C

Abstract

Exploiting the polymerase chain reaction, we have isolated a gene that encodes a putative phosphoinositide-specific phospholipase C (PLC) of the fission yeast Schizosaccharomyces pombe. Inspection of the nucleotide sequence of the gene revealed an open reading frame that can encode a polypeptide of 899 amino acid residues with a calculated molecular mass of 102 kDa. This putative polypeptide contains both the X and Y regions that are conserved among three classes of mammalian PLC, and also contains a presumptive Ca(2+)-binding site (an E-F hand motif). The structure of the putative protein is most similar to that of the delta class of PLC isozymes. To investigate the role of this gene, designated plc1+, gene disruption was carried out by interrupting the coding region with the ura4+ marker. Growth of plc1 cells was temperature-sensitive in rich medium, and cells could not grow in synthetic medium. Expression of the PLC1 gene of Saccharomyces cerevisiae suppressed the growth defect phenotype of plc1- cells, a strong suggestion that the plc1+ gene encodes PLC.