Structural and functional characterization of a single-chain peptide - MHC molecule that modulates both naive and activated CD8 ⁺ T cells

Peptide - MHC (pMHC) multimers, in addition to being tools for tracking and quantifying antigen-specific T cells, can mediate downstream signaling after T-cell receptor engagement. In the absence of costimulation, this can lead to anergy or apoptosis of cognate T cells, a property that could be exploited in the setting of autoimmune disease. Most studies with class I pMHC multimers used noncovalently linked peptides, which can allow unwanted CD8 ⁺ T-cell activation as a result of peptide transfer to cellular MHC molecules. To circumvent this problem, and given the role of self-reactive CD8 ⁺ T cells in the development of type 1 diabetes, we designed a single-chain pMHC complex (scK ^d.IGRP) by using the class I MHC molecule H-2K ^d and a covalently linked peptide derived from islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP _206-214), a well established autoantigen in NOD mice. X-ray diffraction studies revealed that the peptide is presented in the groove of the MHC molecule in canonical fashion, and it was also demonstrated that scK ^d.IGRP tetramers bound specifically to cognate CD8 ⁺ T cells. Tetramer binding induced death of naive T cells and in vitro- and in vivo-differentiated cytotoxic T lymphocytes, and tetramer-treated cytotoxic T lymphocytes showed a diminished IFN-γ response to antigen stimulation. Tetramer accessibility to disease-relevant T cells in vivo was also demonstrated. Our study suggests the potential of single-chain pMHC tetramers as possible therapeutic agents in autoimmune disease. Their ability to affect the fate of naive and activated CD8 ⁺ T cells makes them a potential intervention strategy in early and late stages of disease.