Synthesis of fluorescent lactosylceramide stereoisomers

Abstract

The intracellular distribution of synthetic glycosphingolipids (GSLs) bearing a fluorophore can be monitored in living cells by fluorescence microscopy. We reported previously that variation in the length of the long-chain base and in the structure of the carbohydrate-containing polar head group of (2 S,3 R) (or d- erythro-)-β-lactosylceramide (LacCer) did not alter the mechanism of endocytic uptake from the plasma membrane of various mammalian cell types [Singh, R.D., Puri, V., Valiyaveettil, J.T., Marks, D.L., Bittman, R., Pagano, R.E., 2003. Selective caveolin-1-dependent endocytosis of glycosphingolipids. Mol. Biol. Cell 14, 3254–3265]. To extend our examination of the molecular features in LacCer that are responsible for its uptake by the caveolar-requiring endocytic pathway, we have synthesized the three unnatural stereoisomers [(2 R,3 R)-, (2 S,3 S)-, and (2 R,3 S)] of dipyrromethene difluoride (BODIPY™)-LacCer. These analogues will be used to probe the role of stereochemistry in the long-chain base of LacCer in the mechanism of endocytic uptake.