A two-hybrid antibody micropattern assay reveals specific <i>in cis</i> interactions of MHC I heavy chains at the cell surface.

Abstract

We demonstrate a two-hybrid assay based on antibody micropatterns to study protein-protein interactions at the cell surface of major histocompatibility complex class I (MHC I) proteins. Anti-tag and conformation-specific antibodies are used for individual capture of specific forms of MHC I proteins that allow for location- and conformation-specific analysis by fluorescence microscopy. The assay is used to study the <i>in cis</i> interactions of MHC I proteins at the cell surface under controlled conditions and to define the involved protein conformations. Our results show that homotypic <i>in cis</i> interactions occur exclusively between MHC I free heavy chains, and we identify the dissociation of the light chain from the MHC I protein complex as a condition for MHC I <i>in cis</i> interactions. The functional role of these MHC I protein-protein interactions at the cell surface needs further investigation. We propose future technical developments of our two-hybrid assay for further analysis of MHC I protein-protein interactions.