Role of an ancestral d-bifunctional protein containing two sterol-carrier protein-2 domains in lipid uptake and trafficking in Toxoplasma.

[zellweger syndrome]

The inability to synthesize cholesterol is universal among protozoa . The intracellular pathogen Toxoplasma depends on host lipoprotein-derived cholesterol to replicate in mammalian cells . Mechanisms of cholesterol trafficking in this parasite must be important for delivery to proper organelles . We characterized a unique d-bifunctional protein variant expressed by Toxoplasma consisting of one N-terminal d-3 -hydroxyacyl-CoA dehydrogenase domain fused to two tandem sterol carrier protein-2 (SCP-2 ) domains . This multidomain protein undergoes multiple cleavage steps to release free SCP-2 . The most C-terminal SCP-2 carries a PTS 1 that directs the protein to vesicles before processing . Abrogation of this signal results in SCP-2 accumulation in the cytoplasm . Cholesterol specifically binds to parasite SCP-2 but with 10 -fold lower affinity than phosphatidylcholine . In mammalian cells and Toxoplasma , the two parasite SCP-2 domains promote the circulation of various lipids between organelles and to the surface . Compared with wild-type parasites , TgHAD-2 SCP-2 -transfected parasites replicate faster and show enhanced uptake of cholesterol and oleate , which are incorporated into neutral lipids that accumulate at the basal end of Toxoplasma . This work provides the first evidence that the lipid transfer capability of an ancestral eukaryotic SCP-2 domain can influence the lipid metabolism of an intracellular pathogen to promote its multiplication in mammalian cells .