Andoh T, Yoko T, Matsui Y, Toh A
Department of Plant Sciences, Graduate School of Science, University of Tokyo, Japan.
Yeast. 1995 Feb;11(2):179-85. doi: 10.1002/yea.320110209.
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.
利用聚合酶链反应，我们分离出了一个基因，该基因编码粟酒裂殖酵母的一种假定的磷脂酰肌醇特异性磷脂酶C（PLC）。对该基因核苷酸序列的检查揭示了一个开放阅读框，它可以编码一个由899个氨基酸残基组成的多肽，计算分子量为102 kDa。这个假定的多肽包含在三类哺乳动物PLC中保守的X和Y区域，还包含一个假定的Ca(2+)结合位点（一个E-F手基序）。该假定蛋白质的结构与PLC同工酶的δ类最为相似。为了研究这个命名为plc1+的基因的作用，通过用ura4+标记中断编码区进行了基因破坏。plc1细胞在丰富培养基中的生长对温度敏感，并且在合成培养基中不能生长。酿酒酵母的PLC1基因的表达抑制了plc1-细胞的生长缺陷表型，这有力地表明plc1+基因编码PLC。
