Dev. Cell 23, 573–586 (2012)

The ganglioside GM1 acts as a lipid-based receptor for internalization and traffic of cholera toxin (CT), which is ultimately sorted to the endoplasmic reticulum of host cells. As with other acidic glycosphingolipids, both the extracellular oligosaccharide and the membrane-anchoring ceramide of GM1 have rich structural diversity. Because different GM1 species have different capacities to sort CT to the endoplasmic reticulum, Chinnapen et al. set out to determine whether this was due to the specific ceramide structures. The authors synthesized five different GM1 structures, differing only in the fatty acyl chains in the ceramide domain, and monitored their trafficking in the presence and absence of CT in live cells by fluorescence microscopy. They found that sorting of the different GM1 species from the plasma membrane ultimately to the endoplasmic reticulum or from the plasma membrane to late endosomes is dictated by the ceramide domain. Only the short-chain and the unsaturated species trafficked retrograde into the endoplasmic reticulum, suggesting that these species are the functional CT receptors. Indeed, only these GM1 species led to downstream signaling. Further experiments examining the effect of CT binding on GM1 traffic and of disruption of rafts revealed that a lipid-raft mechanism is most likely involved in GM1 sorting at steps leading into the retrograde endoplasmic reticulum–directed pathway. Therefore, both major tenets of lipid sorting—the 'molecular shape' and 'lipid raft' hypotheses—regulate GM1 and CT traffic.