CD1c presents lipid antigens through a unique dual-presentation mechanism whereby two lipids bind simultaneously in a 2:1 stoichiometry, with bulky gangliosides oriented sideways to protrude laterally through an evolutionarily conserved G'-portal rather than upward through the F'-portal, allowing autoreactive TCR recognition while modulating T cell responses to upright antigens through ligand-dependent conformational changes between open and closed states.

Here we report mass spectrometry analyses of endogenous lipids captured by CD1c when bound to an autoreactive αβTCR. CD1c bound twenty-six lipids with bulky headgroups that could not fit within the tight TCR-CD1c interface. We determined the crystal structures of CD1c presenting several gangliosides, revealing a general mechanism whereby two lipids, rather than one, are bound in the CD1c cleft. Bulky lipids were orientated sideways so that their polar headgroups protruded laterally through a side portal of the CD1c molecule - an evolutionarily conserved structural feature. The sideways presented ganglioside headgroups did not hinder TCR binding and so represent a mechanism that allows autoreactive TCR recognition of CD1c. In addition, ex vivo studies showed sideways presented gangliosides could also represent TCR recognition determinants. These findings reveal a general mechanism whereby CD1c simultaneously presents two lipid antigens from the top and side of its cleft that differs markedly from other antigen-presenting molecules.