An altered invariant chain protein with an antigenic peptide in place of CLIP forms SDS-stable complexes with class II αβ dimers and facilitates highly efficient peptide loading

We report an experimental system for abundant expression of specific peptide-class II complexes in vivo and in vitro. We have constructed a cassette which allows for the replacement of the CLIP region of invariant chain (li) with an antigenic peptide. In fibroblasts expressing an altered ii protein, in which CLIP has been replaced with peptide 52-68 from the class II I-E alpha chain (pE alpha), pE alpha-I-A(b) complexes are formed with high efficiency. This peptide loading occurs in the endoplasmic reticulum (ER) when the Ii:pE alpha fusion protein associates with the I-A(b) alpha and beta chains. The trimeric complexes of Ii:pE alpha and I-A(b) molecules are stable in SDS and can be detected by the pE alpha-I-A(b)-specific mAb, YAe, indicating that pE alpha is bound in the class II groove in the context of full-length Ii. These data strongly suggest that the CLIP region of intact Ii prevents peptide loading in the ER by binding in the peptide binding groove of newly synthesized class II alpha beta dimers.