Growth of a mutant defective in a putative phosphoinositide-specific phospholipase C of Schizosaccharomyces pombe is restored by low concentrations of phosphate and inositol

Abstract

A mutant (plc1-1) of Schizosaccharomyces pombe unable to grow on a minimal medium containing high amounts of phosphate was selected. On yeast-extract agar its growth is temperature sensitive. Tests in liquid synthetic medium show that growth of the mutant is partially restored by lowering the phosphate and inositol concentrations in the growth medium. The growth defect is fully suppressed by a plasmid encoding a putative protein having the structural features of phosphoinositide-specific phospholipases C (PI-PLC). This protein, of 899 amino-acids, contains the characteristic X and Y domains found in all PI-PLCs of higher and lower eucaryotes and reveals, in addition, an EF-hand motif (putative Ca(2+)-binding site). Like the corresponding enzyme from Saccharomyces cerevisiae, the S. pombe PI-PLC is most similar to the delta form of PI-PLC isoenzymes. The cloned gene integrates at the plc1 site indicating that plc1 codes for a putative PI-PLC. Plc1 physically maps on the left arm of chromosome II between rad11 and mei3.

A gene encoding phosphatidyl inositol-specific phospholipase C from Cryphonectria parasitica modulates the lac1 expression

“…PLC genes have been identiWed in diverse eukaryotic organisms other than mammals, including the slime mold (Dictyostelium discoideum) (Drayer and Van Haastert, 1992) and yeast (Bennett et al, 1998;Fankhauser et al, 1995;Yoko-o et al, 1993). In addition, there is growing evidence to implicate phosphoinositides in the signal transduction pathway of Wlamentous fungi (Gadd, 1995).…”

A Schizosaccharomyces pombe gene, ksg1, that shows structural homology to the human phosphoinositide-dependent protein kinase PDK1, is essential for growth, mating and sporulation

“…It has been shown that the percentage of phosphatidylinositol present in phospholipids strongly depends on the inositol concentration in the growth medium (Fernandez et al 1986). Genes encoding phospholipase C and phosphatidylinositol-speci®c PI 3-kinase have been identi®ed (Andoh et al 1995;Fankhauser et al 1995a;Takegawa et al 1995). Phospholipase C interacts with the 14-3-3 protein encoded by rad24 and is involved in the control of resistance to UV irradiation (Andoh et al 1998).…”

Molecular Cloning and Expression of a Mouse Thiamin Pyrophosphokinase cDNA

“…The deduced amino acid sequence of the S. pombe TPK gene (tnr3) (17) was used to conduct a text-based search of the mouse EST database at the National Center for Biotechnology Information. The sequence information of the obtained mouse clone AA981202 allowed the synthesis of three specific primers, F1 (5Ј-TGACCAAATCATGGC-CTCTGTG), R1 (5Ј-TTCATGCTGAGCAACGAACCAG), and R2 (5Ј-TTAGCTCTTGATGGCCATGGT).…”

“…4). The amino acid sequence of mTPK1 showed 39% identity with the carboxyl-terminal half of tnr3, which appears as a fusion protein (569 amino acids) with unknown function at the amino-terminal half (17). The mTPK1 protein was also similar to THI80 and pTPK1 over their full lengths (31% of amino acid identity).…”

“…Recently, the mammalian thiamin transport protein, THTR-1, was identified as the gene mutated in thiamin-responsive megaloblastic anemia (Online Mendelian Inheritance in Man number: 249270) (10 -12). However, although the enzyme has been purified from several mammalian sources (13)(14)(15), the nucleotide sequences of TPK have been determined only for the microorganisms (16,17) Saccharomyces cerevisiae (THI80), Schizosaccharomyces pombe (tnr3), and Paracoccus denitrificans (pTPK1). The isolation of the cDNA or genomic DNA for TPK from mammalian cells is indispensable for the elucidation of thiamin metabolism and its regulation in higher animals.…”