The transcription factor Duxbl mediates elimination of pre-T cells that fail β-selection

Abstract

T cell development is critically dependent on successful rearrangement of antigen-receptor chains. At the β-selection checkpoint, only cells with a functional rearrangement continue in development. However, how nonselected T cells proceed in their dead-end fate is not clear. We identified low CD27 expression to mark pre-T cells that have failed to rearrange their β-chain. Expression profiling and single-cell transcriptome clustering identified a developmental trajectory through β-selection and revealed specific expression of the transcription factor Duxbl at a stage of high recombination activity before β-selection. Conditional transgenic expression of Duxbl resulted in a developmental block at the DN3-to-DN4 transition due to reduced proliferation and enhanced apoptosis, whereas RNA silencing of Duxbl led to a decrease in apoptosis. Transcriptome analysis linked Duxbl to elevated expression of the apoptosis-inducing Oas/RNaseL pathway. RNaseL deficiency or sustained Bcl2 expression led to a partial rescue of cells in Duxbl transgenic mice. These findings identify Duxbl as a regulator of β-selection by inducing apoptosis in cells with a nonfunctional rearrangement.

Developing T cells form an immunological synapse for passage through the β-selection checkpoint

“…To compare PMA treatment with pre-TCR signaling, we established a system in which cells preferentially progressed either though the β-selection checkpoint via PMA signaling (DN3a cells that had not yet expressed the pre-TCR) or only via signaling through the pre-TCR (a mixed population of DN3a cells both expressing and not expressing TCRβ that were not treated with PMA). Consistent with recently published work ( Klein et al, 2019 ), DN3a cells that expressed cell surface TCRβ differentiated faster and proliferated more than those that did not express TCRβ, compatible with pre-TCR being a prerequisite for progression beyond the β-selection checkpoint. We used this system to discriminate between a role for Notch and CXCR4 in immunological synapse assembly compared with downstream signaling.…”

“…Consistent with recently published work (Klein et al, 2019), DN3a cells that expressed cell surface TCRβ differentiated faster and proliferated more than those that did not express TCRβ, compatible with the pre-TCR as being a prerequisite for progression beyond the β-selection checkpoint. We used this system to discriminate between a role for Notch and CXCR4 in immunological synapse assembly compared with downstream signaling.…”

“…To avoid confounding the results with cells that had already assembled an immunological synapse, we sorted DN3a cells for the presence or absence of surface TCRβ. As previously published, the cell surface expression of TCRβ dramatically alters subsequent fate of the cell populations ( Klein et al, 2019 ). After 5 d of coculturing on OP9-DL1, TCRβ + cells yielded more than double the cells yielded by TCRβ − cells, with commensurate differences in CFSE dilution, indicating higher cell division rates ( Fig.…”

DUX4 Role in Normal Physiology and in FSHD Muscular Dystrophy

“…In thymus, successful rearrangement of the T cell receptor β-chain is required for proper T cell development and cells with a non-functional rearrangement undergo apoptosis. Intriguingly, Duxbl, another DUX4 homolog in mouse, is selectively expressed just before β-selection and plays an important role in elimination of cells that have failed rearrangement of the T cell receptor β-chain [ 38 , 39 ].…”

Single‐cell transcriptomics uncovers an instructive T‐cell receptor role in adult γδ T‐cell lineage commitment

“…However, individual T cells within a defined developmental stage might still display some heterogeneity reflecting different developmental potential, an issue that cannot be unveiled by "bulk" transcriptomic studies (Mingueneau et al, 2013;Roels et al, 2020). In contrast, single-cell RNA sequencing (scRNAseq) permits to analyze genome-wide RNA expression at single-cell levels and to determine the degree of transcriptomics heterogeneity of a given T-cell precursor population (Klein et al, 2019, Oh et al, 2021, Rothenberg, 2021, Sagar et al, 2020. Here, we used scRNAseq to analyze the developmental bifurcation leading to ab and cd T cells in the thymus of adult wild-type (WT) and LAT-deficient mice.…”