The Initial Step of the Glycerolipid Pathway IDENTIFICATION OF GLYCEROL 3-PHOSPHATE/DIHYDROXYACETONE PHOSPHATE DUAL SUBSTRATE ACYLTRANSFERASES IN SACCHAROMYCES CEREVISIAE *

The initial step of phospholipid biosynthesis in yeast is carried out through the acylation of glycerol 3-phos-phate (G-3-P) and dihydroxyacetone phosphate by stereospecific sn -1 acyltransferases. Here we report the identification of two key fatty acyltransferases of the glycerolipid biosynthesis pathway in Saccharomyces cerevisiae . Disruption of the open reading frame YBL011w , corresponding to a gene previously identified as a choline transporter suppressor ( SCT1 ), resulted in a substantial decrease of total cellular G-3-P acyltransferase activity. A yeast strain disrupted at the open reading frame YKR067w , which encodes a protein closely related to Sct1p, also exhibited a dramatic reduction in G-3-P acyltransferase activity. Molecular characterizations of the genes revealed that a missense mutation in YKR067w accounted for a defect in the activities of the G-3-P acyltransferase in the yeast mutant strain TTA1. Heterologous expression of YKR067w in Escherichia coli further confirmed its enzyme activity. These results indicate that YKR067w and YBL011w , designated herein as GAT1 and GAT2(SCT1) , respectively, are yeast G-3-P acyltransferase genes. Furthermore, biochemical results are presented to show that both Gat1p