Use of fluorescent sphingolipid analogs to study lipid transport along the endocytic pathway

Sphingolipids (SLs) are concentrated at the plasma membrane where they play important roles in cell cell communication, host pathogen interactions. and cell signaling events. Our laboratory has used fluorescent SL analogs and SI.-binding toxins to elucidate mechanisms by which SLs are internalized by endocytosis and subsequently sorted and transported to various intracellular compartments. These studies have relied on the use of temperature shift protocols, co-localization studies with compartment-specific markers, selective biochemical treatments that inhibit specific endocytic mechanisms. kind the expression of dominant negative proteins (e.g., rabs) that block specific steps in transport. These methods are presented here so that they can he utilized by others for the study of endocytic trafficking of lipids and other molecules. (c) 2005 Elsevier Inc. All rights reserved.