Abstract

Peptide exchange technologies are essential for the generation of pMHC-multimer libraries used to probe diverse, polyclonal TCR repertoires in various settings. Here, using the molecular chaperone TAPBPR, we develop a robust method for the capture of stable, empty MHC-I molecules comprising murine H2 and human HLA alleles, which can be readily tetramerized and loaded with peptides of choice in a high-throughput manner. Alternatively, catalytic amounts of TAPBPR can be used to exchange placeholder peptides with high affinity peptides of interest. Using the same system, we describe high throughput assays to validate binding of multiple candidate peptides on empty MHC-I/TAPBPR complexes. Combined with tetramer-barcoding via a multi-modal cellular indexing technology, ECCITE-seq, our approach allows a combined analysis of TCR repertoires and other T cell transcription profiles together with their cognate antigen specificities in a single experiment. The new approach allows TCR/pMHC interactions to be interrogated easily at large scale.

Peptide-MHC (pMHC) tetramers are important tools for probing T cell repertoire and adaptive immune responses. Here the authors use a molecular chaperone, TAPBPR, to develop a high-throughput, multiplexible platform for pMHC tetramer generation to facilitate simultaneous assessments of T cell repertoire/antigen specificity and transcriptome.

Details

Title
High throughput pMHC-I tetramer library production using chaperone-mediated peptide exchange
Author
Overall, Sarah A 1 ; Toor, Jugmohit S 1 ; Hao, Stephanie 2 ; Yarmarkovich, Mark 3   VIAFID ORCID Logo  ; Sara M O’Rourke 1 ; Morozov, Giora I 1 ; Nguyen, Son 4   VIAFID ORCID Logo  ; Sada, Japp Alberto 4   VIAFID ORCID Logo  ; Gonzalez, Nicolas 1 ; Moschidi Danai 1 ; Betts, Michael R 4 ; Maris, John M 3 ; Smibert, Peter 2   VIAFID ORCID Logo  ; Sgourakis, Nikolaos G 1 

 University of California Santa Cruz, Department of Chemistry and Biochemistry, Santa Cruz, USA (GRID:grid.205975.c) (ISNI:0000 0001 0740 6917) 
 New York Genome Center, Technology Innovation Lab, New York, USA (GRID:grid.429884.b) 
 University of Pennsylvania, Division of Oncology, Center for Childhood Cancer Research, Children’s Hospital of Philadelphia and Department of Pediatrics, Perelman School of Medicine, Philadelphia, USA (GRID:grid.25879.31) (ISNI:0000 0004 1936 8972) 
 University of Pennsylvania, Department of Microbiology, Perelman School of Medicine, Philadelphia, USA (GRID:grid.25879.31) (ISNI:0000 0004 1936 8972) 
Publication year
2020
Publication date
2020
Publisher
Nature Publishing Group
e-ISSN
20411723
Source type
Scholarly Journal
Language of publication
English
ProQuest document ID
2392415961
Copyright
© The Author(s) 2020. This work is published under http://creativecommons.org/licenses/by/4.0/ (the “License”). Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.