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Introduction: While most transcripts arising from the human T Cell Receptor locus reflect fully rearranged genes, several germline transcripts have been identified. We describe a new germline transcript arising from the human TCRB locus. Methods: cDNA sequencing, promoter, and gene expression analyses were used to characterize the new transcript. Results: The new germline transcript encoded by the human TCRB locus consists of a new exon of 103bp, which we named TRBX1 (X1), spliced with the first exon of gene segments C ss 1 or C ss 2. X1 is located upstream of gene segment D ss 1 and is therefore deleted from a V-DJ rearranged TCRB locus. The X1-C ss transcripts do not appear to code for a protein. We define their transcription start and minimal promoter. These transcripts are found in populations of mature T lymphocytes from blood or tissues and in T cell clones with a monoallelic TCRB rearrangement. In immature thymocytes, they are already detectable in CD1a(-)CD34(+)CD4(-)CD8(-) cells, therefore before completion of the TCRB rearrangements. Conclusions: The X1 promoter appears to be the ortholog of the mouse pre-D ss 1 promoter (PD ss 1). Like PD ss 1, its activation is regulated by E ss in T cells and might facilitate the TCRB rearrangement process by contributing to the accessibility of the D ss 1 locus.

Regulatory T lymphocytes (Tregs) are a subset of CD4+ T cells specialised in the inhibition of immune responses. Tregs are required for the maintenance of self-tolerance, but in tumours they play a detrimental role by suppressing T lymphocytes directed against cancer cells. Although suppression mechanisms are not fully elucidated yet, Tregs inhibit other immune cells at least in part through the production of active TGF-β1. Most hematopoietic cells, including Tregs, secrete TGF-β1 under a latent form, in which the mature cytokine is kept inactive by association with the so-called Latency-Associated Peptide (LAP). Stimulation of the T Cell Receptor (TCR) induces expression of transmembrane protein GARP on the surface of Tregs, but not other T lymphocytes. GARP binds and presents latent TGF-β1, and allows release of mature TGF-β1 from the LAP close to the Treg surface. GARP may therefore play an important role in Treg suppression through control of active TGF-β1 production. The objectives of our work were twofold. First, we sought to determine whether GARP, in addition to activating latent TGF-β1, also controls earlier steps of the TGF-β1 production pathway in human Tregs. We found that GARP favours the cleavage of the pro-TGF-β1 precursor, and increases secretion of latent TGF-β1. We also found that stimulated Tregs secrete latent TGF-β1 as complexes containing GARP. GARP/TGF-β1 complexes represent a novel form of secreted TGF-β1, with as yet unknown biological function. Second, we sought to identify transcriptional and/or post-transcriptional mechanisms that regulate GARP expression, and hence TGF-β1 production, in human T cells. We showed that GARP levels are regulated post-transcriptionally by miR-142-3p, miR-181a, b, c, d and miR-185. High expression of these miRNAs in non-regulatory T cells may prevent appearance of the GARP protein even in the presence of high GARP mRNA levels. In addition, we found that GARP expression is inhibited at the transcriptional level by TGF-β1 signals. Considering previous data and our own, we propose the following model. TCR stimulation of human Tregs induces GARP expression. This increases latent TGF-β1 production, induces secretion of GARP/TGF-β1 complexes, presentation of latent TGF-β1 at the Treg surface, and production of active TGF-β1. Autocrine TGF-β1 signals, in turn, repress transcription of the GARP gene. Downregulation of GARP by autocrine TGF-β1 in Tregs represents a negative feedback mechanism resulting in the return to a resting state, characterised by the absence of GARP, reduced latent TGF-β1 secretion and shut down of active TGF-β1 production. (BIFA - Sciences biomédicales et pharmaceutiques) -- UCL, 2013