Mammalian phosphatidylinositol transfer proteins (PITPs) α and β, which share 77% identity, have been shown to exhibit distinct lipid-transfer activities. In addition to transferring phosphatidylinositol (PI) and phosphatidylcholine (PC), PITPβ has been shown to transfer sphingomyelin (SM), and this has led to the suggestion that PITPβ is important for the regulation of SM metabolism. In the present study, we have analysed the ability of human PITPβ to transfer and regulate the metabolism of cellular SM. We report that, in vitro, the two PITP isoforms were comparable in mediating PI, PC or SM transfer. Using permeabilized HL-60 cells as the donor compartment, both PITP isoforms efficiently transferred PI and PC, and were slightly active towards SM, with the activity of PITPβ being slightly greater. To identify which cellular lipids were selected by PITPs, PITPα and PITPβ were exposed to permeabilized HL-60 cells, and subsequently repurified and analysed for their bound lipids. Both PITPs were able to select only PI and PC, but not SM. SM synthesis takes place at the Golgi, and PITPβ was shown to localize in that compartment. To examine the role of PITPβ in SM biosynthesis, Golgi membranes were used. Purified Golgi membranes had lost their endogenous PITPβ, but were able to recruit PITPβ when added exogenously. However, PITPβ did not enhance the activities of either SM synthase or glucosylceramide synthase. Further analysis in COS-7 cells overexpressing PITPβ showed no effects on (a) SM and glucosylceramide biosynthesis, (b) diacylglycerol or ceramide levels, (c) SM transport from the Golgi to the plasma membrane, or (d) resynthesis of SM after exogenous sphingomyelinase treatment. Altogether, these observations do not support the suggestion that PITPβ participates in the transfer of SM, the regulation of SM biosynthesis or its intracellular trafficking.
Phosphatidylinositol transfer protein β displays minimal sphingomyelin transfer activity and is not required for biosynthesis and trafficking of sphingomyelin
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Bruno SÉGUI, Victoria ALLEN-BAUME, Shamshad COCKCROFT; Phosphatidylinositol transfer protein β displays minimal sphingomyelin transfer activity and is not required for biosynthesis and trafficking of sphingomyelin. Biochem J 15 August 2002; 366 (1): 23–34. doi: https://doi.org/10.1042/bj20020317
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