Clifford Lingwood
University of Toronto, Canada
Title: ABC cassette transporters regulate glycosphingolipid biosynthesis
Biography
Biography: Clifford Lingwood
Abstract
ABC cassette transporters differentially control GSL synthesis. The precursor for the synthesis of most complex glycosphingolipids (GSLs) is glucosylceramide (GlcCer) made on the outer Golgi membrane. The mechanism by which GlcCer is flipped into the Golgi has remained a mystery for>40yeras. Based on the use of MDR1(ABCB1) inhibitors, we proposed a role for this multidrug resistance protein. We have now used a newly synthesized photolabile GlcCer crosslinker to identify GlcCer binding proteins in microsomes, using proteomics/mass spectrometry. Three ATP binding ABC cassette transporters were thus identified, ABCB10, ABCB4 and ABCA3. Using SiRNA knockdown, these candidate GlcCer flippases were reduced in DU145 cells. The loss of ABC cassette proteins had a differential effect on selective GSLs. Some were increased e.g. lactosyl ceramide, while others reduced. In each case of ABC cassette knockdown, GalCer was increased. Moreover, MDR1 siRNA knockdown reduced overall GSL content by>50% according to cell line. These studies provide new insights into the complex regulation of GSL biosynthesis by precursor supply, a new basis for the link between cancer/multidrug resistance and GSL biosynthesis, suggest a new link between GlcCer and GalCer-based GSLs and indicate that multiple ABC transporters can transport GlcCer into the Golgi to generate LacCer pools used for the synthesis of different GSLs. Different LacCer pools could provide a partial basis for the biosynthesis of different GSLs within different Golgi regions (figure).
References
1. Mahfoud R, Manis A, Binnington B, Ackerley C, Lingwood CA (2010) A major fraction of glycosphingolipids in model and cellular cholesterol containing membranes are undetectable by their binding proteins. J Biol Chem 285: 36049-36059.
2. Kamani M, Mylvaganam M, Tian R, Binnington B, Lingwood C (2011) Adamantyl glycosphingolipids provide a new approach to the selective regulation of cellular glycosphingolipid metabolism. J Biol Chem 286: 21413-21426.
3. Lingwood D, Binnington B, Róg T, Vattulainen I, Grzybek M, et al. (2011) Cholesterol modulates glycolipid conformation and receptor activity. Nature Chem Biol 7: 260-262.
4. Kamani MA, Provencal P, Boutin M, Pacienza N, Fan X, et al. (2016) Glycosphingolipid storage in Fabry mice extends beyond globotriaosylceramide and is affected by ABCB1 depletion. Future Sci OA 2: FSO147.
5. Lingwood CA (2011) Glycosphingolipid Functions. Cold Spring Harb Perspect Biol Biology of Lipids: 149-174.