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. 2019 Oct 1;10:4451. doi: 10.1038/s41467-019-12441-w

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© The Author(s) 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 2 — Conversion of a weak TCR into a dominant TCR by replacement of 14 variable region framework residues. a Schematic representation of the retroviral vector used for TCR expression studies. TCR α and β chain expression was determined using antibodies specific for the V5 and myc epitopes, respectively. Transduction efficiency was determined using antibodies specific for murine CD19. b Representative example of n = 4 independent experiments showing human Jurkat cells (expressing an endogenous TCR) transduced with a dominant (Dom) TCR (TRAV38-2/TRBV7-8) or three different weak TCRs: weak 1 (TRAV13-2/TRBV7-3), weak 2 (TRAV23/TRBV7-9) or weak 3 TCR (TRAV29/TRBV2). Top panel: CD19 expression levels. Middle panel: TCR α and β chain expression levels on gated CD19^high cells. Bottom panel: TCR α and β chain expression levels on gated CD19^intermediate cells. c Pooled data (means ± SEM) showing TCR α and β chain expression levels normalized to the Dom TCR. n = 4 independent experiments. Top panel: gated on CD19^high cells. Bottom panel: gated on CD19^intermediate cells. * P < 0.05 (Mann–Whitney U test) for all comparisons between the Dom TCR α chain and the weak TCR α chains and for all comparisons between the Dom TCR β chain and the weak TCR β chains. MFI, median fluorescence intensity. d Top panel: introduction of the 14 residues indicated in Fig. 1e into the weak 1 TCR (TRAV13-2/TRBV7-3) generated the weak → dom TCR with enhanced α/β expression on the cell surface. Bottom panel: replacement of the 14 residues in the Dom TCR (TRAV38-2/TRBV7-8) with the equivalent residues in the weak 1 TCR (TRAV13-2/TRBV7-3) generated the dom → weak TCR with undetectable α/β expression on the cell surface. TCR constructs were transduced into Jurkat cells expressing an endogenous TCR. Data are representative of four independent experiments. e Pooled data (means ± SEM) showing TCR α and β chain expression levels normalized to the corresponding unmodified TCRs. n = 4 independent experiments. *P < 0.05 (Mann–Whitney U test) for all comparisons between the modified TCRs and the corresponding unmodified TCRs. MFI median fluorescence intensity. Vα variable alpha, Vβ variable beta