Inefficient V(D)J recombination underlies monogenic T cell receptor β expression

Abstract

The assembly of T cell receptor (TCR) and immunoglobulin (Ig) genes by V(D)J recombination generates the antigen receptor (AgR) diversity that is vital for adaptive immunity. At most AgR loci, V(D)J recombination is regulated so that only one allele assembles a functional gene, ensuring that nearly every T and B cell expresses a single type, or specificity, of AgR. The genomic organizations of some AgR loci permit the assembly and expression of two distinct genes on each allele; however, this is prevented by undetermined mechanisms. We show that the poor qualities of recombination signal sequences (RSSs) flanking Vβ gene segments suppress the assembly and expression of two distinct TCRβ genes from a single allele. Our data demonstrate that an intrinsic genetic mechanism that stochastically limits Vβ recombination efficiency governs monogenic TCRβ expression, thereby restraining the expression of multiple AgRs on αβ T cells.

Keywords: V(D)J recombination; allelic exclusion; lymphocyte development; monogenic expression; recombination signal sequence.

Fig. 1.

Increased frequency of dual-TCRβ⁺ cells and altered Vβ repertoire in mice with two Vβ RSS replacements on the same allele. (A) Schematic of the TCRβ locus and relative positions of Vβ, Dβ, and Jβ segments, Cβ exons, and the Eβ enhancer. (B and C) Representative plots (B) and quantification (C) of SP thymocytes expressing both V2⁺ and V31⁺ TCRβ chains. (D–F) Representative plots (D) and quantification of SP thymocytes expressing V2⁺ (E) or V31⁺ (F) TCRβ chains. n ≥ 4 mice per group; one-way ANOVA, Dunnett’s multiple posttests comparing each RSS mutant to WT; ***P < 0.001, ****P < 0.0001. Data in B–F are compiled from five experiments.

Fig. 2.

Expression of two different TCRβ chains from the V2^RV31^R allele. (A and B) Representative plots (A) and quantification (B) of SP thymocytes expressing both V2⁺ and V31⁺ TCRβ chains (n ≥ 6 mice per group; one-way ANOVA, Tukey’s multiple posttests; ns, not significant; ****P < 0.0001). (C) Schematic of the sense strand and truncation of the V31 region of the V2^R31Δ allele, with the V31 RSS indicated in blue. (D) Depiction of the recombination events that could result in two TCRβ chains expressed from one allele. RSSs are indicated as triangles. Data in A and B are compiled from four experiments.