Cholera toxin causes diarrheal disease by binding ganglioside GM1 on the apical membrane of polarized intestinal epithelial cells and trafficking retrograde through sorting endosomes, the trans-Golgi network (TGN), and into the endoplasmic reticulum. A fraction of toxin also moves from endosomes across the cell to the basolateral plasma membrane by transcytosis, thus breeching the intestinal barrier. Here we find that sorting of cholera toxin into this transcytotic pathway bypasses retrograde transport to the TGN. We also find that GM1 sphingolipids can traffic from apical to basolateral membranes by transcytosis in the absence of toxin binding but only if the GM1 species contain cis-unsaturated or short acyl chains in the ceramide domain. We found previously that the same GM1 species are needed to efficiently traffic retrograde into the TGN and endoplasmic reticulum and into the recycling endosome, implicating a shared mechanism of action for sorting by lipid shape among these pathways. Background: Mechanisms for intracellular lipid sorting remain poorly understood. Results: Polarized epithelial cells sort ganglioside GM1, the receptor for cholera toxin, into distinct retrograde and transcytotic pathways, provided that GM1 contains ceramide domains with short or unsaturated fatty acid chains. Conclusion: Sphingolipid sorting depends on ceramide structure, implicating a mechanism for lipid sorting by lipid shape. Significance: The results identify a lipid-sorting pathway across epithelial barriers with clinical applications.