Production of MR1 Tetramers Loaded with Microbial Ligands
In lieu of peptides, the monomorphic MHC-I-like molecule MR1 presents small molecule antigens to stimulate a subset of αβ T cells known as mucosal-associated (semi-) invariant T (MAIT) cells or, more broadly, MR1-restricted (MR1T) cells. The MR1 ligands identified to date are limited to derivatives and intermediates of the riboflavin and folate biosynthesis pathways and their presentation is therefore thought to be an indicator of infection by microbial species that can synthesize riboflavin. MAIT cells have, in recent years, been studied and isolated using a tetrameric reagent of recombinant MR1 loaded with the canonical ligand 5-OP-RU due to its potency toward MAIT clones. However, new evidence has shown that the repertoire of MR1 ligands is much more diverse than previously appreciated and, consistent with this, that the 5-OP-RU tetramer does not bind all MR1T cells. To study MR1-restricted T cell clones in the context of unique bacterial infection, we have generated a tetramer of MR1 loaded with diverse microbial antigens. The production of this reagent is detailed in this chapter.
Key wordsMR1 MAIT Recombinant protein Microbes Mucosal immunity T Cell Receptor MR1T
We would like to acknowledge the Bill and Melinda Gates Foundation for funding this protocol development, grant # OPP1131709, and the T32 Training Grant to the University of Chicago: GM007183. We thank members of the Lewinsohn Lab at Oregon Health and Science University, Dr. Melanie Harriff, Dr. Adrienne Luoma, Dr. Sobhan Roy, and Dr. Caitlin Castro for helpful discussions during protocol development.
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