Cholesterol inhibits TCR signaling by directly restricting TCR-CD3 core tunnel motility

Cholesterol molecules specifically bind to the resting alpha beta TCR to inhibit cytoplasmic CD3 zeta ITAM phosphorylation through sequestering the TCR-CD3 complex in an inactive conformation. The mechanisms of cholesterol-mediated inhibition of TCR-CD3 and its activation remain unclear Here, we present cryoelectron microscopy structures of cholesterol- and cholesterol sulfate (CS)-inhibited TCR-CD3 complexes and an auto-active TCR-CD3 variant. The structures reveal that cholesterol molecules act like a latch to lock CD3 zeta into an inactive conformation in the membrane. Mutations impairing binding of cholesterol molecules to the tunnel result in the movement of the proximal C terminus of the CD3 zeta transmembrane helix, thereby activating the TCR-CD3 complex in human cells. Together, our data reveal the structural basis of TCR inhibition by cholesterol, illustrate how the cholesterol-binding tunnel is allosterically coupled to TCR triggering, and lay a foundation for the development of immunotherapies through directly targeting the TCR-CD3 complex.

Automated summary

In this study, cryoelectron microscopy structures of cholesterol- and cholesterol sulfate (CS)-inhibited TCR-CD3 complexes and an auto-active TCR-CD3 variant are presented. The structures show that cholesterol molecules act as a latch to lock CD3ζ into an inactive conformation in the membrane. Mutations disrupting the binding of cholesterol molecules to the tunnel result in the activation of the TCR-CD3 complex in human cells.