The gene encoding the phosphatidylinositol transfer protein is essential for cell growth

Abstract

Phosphatidylinositol transfer proteins (PI-TPs) catalyze the transfer of phosphatidylinositol and phosphatidylcholine between membranes in vitro. However, the in vivo function of these proteins is unknown. In this paper, we use a combined biochemical and genetic approach to determine the importance of PI-TP in vivo. An oligonucleotide based on the amino-terminal sequence of the PI-TP from Saccharomyces cerevisiae was used to screen a yeast genomic library for the gene encoding PI-TP (PIT1 gene). Positive clones showed overproduction of transfer activities and transfer protein in the 100,000 x g supernatants. The 5' terminus of the PIT1 gene correlates with the predicted codons for residues 3-30 of the determined protein sequence. A putative intron is located between the codons for residues 2 and 3 of the protein sequence. The codons for the first two amino acids of the protein and the presumptive initiation methionine precede the intron. Tetrad analysis of a heterozygous diploid (PIT1/pit1::LEU2) revealed that the PIT1 gene is essential for cell growth. Nonviable spores could be rescued by transformation of the above diploid prior to sporulation, with a plasmid-borne copy of the wild type gene.