The initial step of glycerolipid metabolism in Leishmania major promastigotes involves a single glycerol-3-phosphate acyltransferase enzyme important for the synthesis of triacylglycerol but not essential for virulence.
The synthesis of the major phospholipids, including those that play an essential role in Leishmania virulence, initiates with the acylation of glycerol-3-phosphate and dihydroxyacetonephosphate at the sn-1 position by glycerol-3-phosphate and dihydroxyacetonephosphate acyltransferases respectively. In this study, we show that Leishmania major promastigotes express a single glycerol-3-phosphate acyltransferase activity important for triacylglycerol synthesis but not essential for virulence. The encoding gene, LmGAT, expressed in yeast results in full complementation of the lethality of a mutant, gat1Deltagat2Delta, lacking glycerol-3-phosphate activity. Biochemical analyses revealed that LmGAT is a low-affinity glycerol-3-phosphate acyltransferase and exhibits higher specific activity with unsaturated long fatty acyl-CoA donors. A L. major null mutant, Deltalmgat/Deltalmgat, was created and a thorough analysis of its lipid composition was performed. Deletion of LmGAT resulted in a complete loss of Leishmania glycerol-3-phosphate acyltransferase activity and a major reduction in triacylglycerol synthesis. Consistent with the specificity of LmGAT for glycerol-3-phosphate but not dihydroxyacetonephosphate, Deltalmgat/Deltalmgat mutant expressed normal levels of the ether-lipid derivatives and virulence factors, lipophosphoglycan and GPI-anchored proteins, gp63, and its virulence was not affected in mice.
Zufferey
Rachel
R
Center for Microbial Pathogenesis, University of Connecticut Health Center, 263 Farmington Ave, Farmington, CT 06030, USA.
Mamoun
Choukri Ben
CB
D000595:Amino Acid Sequence; D000818:Animals; D004099:Dihydroxyacetone Phosphate / Q000378:metabolism; D017125:Genes, Protozoan; D005989:Glycerides / Q000378:metabolism*; D005992:Glycerol-3-Phosphate O-Acyltransferase / Q000235:genetics / Q000378:metabolism*; D005994:Glycerophosphates / Q000378:metabolism; D006028:Glycosphingolipids / Q000378:metabolism; D007700:Kinetics; D018320:Leishmania major / Q000378:metabolism* / Q000472:pathogenicity* / Q000502:physiology; D016773:Leishmaniasis, Cutaneous / Q000469:parasitology; D008666:Metalloendopeptidases / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D013379:Substrate Specificity; D014280:Triglycerides / Q000096:biosynthesis*; D014774:Virulence / Q000502:physiology
D005989:Glycerides; D005994:Glycerophosphates; D006028:Glycosphingolipids; D014280:Triglycerides; C008290:lipophosphonoglycan; D004099:Dihydroxyacetone Phosphate; C029620:alpha-glycerophosphoric acid; D005992:Glycerol-3-Phosphate O-Acyltransferase; D008666:Metalloendopeptidases; C055146:glycoprotein gp63, Leishmania