Phosphatidylethanolamine (PtdEtn) is synthesized by multiple pathways located in different subcellular compartments in yeast. Strains defective in the synthesis of PtdEtn via phosphatidylserine (PtdSer) synthase/decarboxylase are auxotrophic for ethanolamine, which must be transported into the cell and converted to phospholipid by the CDP-Etn dependent Kennedy pathway. We now demonstrate that yeast strains with psd1 psd2 mutations, devoid of PtdSer decarboxylases, import and acylate exogenous lyso-PtdEtn. Lyso-PtdEtn supports growth and replaces the mitochondrial pool of PtdEtn much more efficiently than, and independently of PtdEtn derived from the Kennedy pathway. Deletion of both the PtdSer decarboxylase and Kennedy pathways yields a strain that is a stringent lyso-PtdEtn auxotroph. Evidence for the specific uptake of lyso-PtdEtn by yeast comes from analysis of strains harboring deletions of the aminophospholipid translocating P-type ATPases (APLTs). Elimination of the APLTs, Dnf1p and Dnf2p, or their non-catalytic beta-subunit, Lem3p, blocked the import of radiolabeled lyso- PtdEtn, and resulted in growth inhibition of lyso-PtdEtn auxotrophs. In cell extracts, lyso-PtdEtn is rapidly converted to PtdEtn by an acyl-CoA dependent acyltransferase. These results now provide 1) an assay for APLT function based on an auxotrophic phenotype, 2) direct demonstration of APLT action on a physiologically relevant substrate, and 3) a genetic screen aimed at finding additional components that mediate the internalization, trafficking, and acylation of exogenous lyso-phospholipids.
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