BTLA and PD-1 signals attenuate TCR-mediated gene expression
T cell coinhibitory immune checkpoints, such as PD-1 or BTLA, are bona fide targets in cancer therapy. Here we used a human T cell reporter line to measure transcriptome changes mediated by PD-1 and BTLA induced signalling. TCR/CD3 stimulation resulted in the upregulation of a large number of genes but also repressed a similar number of transcripts. PD-1 and BTLA signals attenuated transcriptomics changes mediated by TCR/CD3 signalling: upregulated genes tended to be suppressed and the expression of a significant number of downregulated genes was higher when PD-1 or BTLA signalling took place. BTLA was a significantly stronger attenuator of TCR/CD3 induced transcriptome changes than PD-1. A strong overlap between genes that were regulated indicated quantitative rather than qualitative differences between these receptors. In line with their function as attenuator of TCR/CD3 mediated changes we found strongly regulated genes to be prime targets of PD-1 and BTLA signalling. Overall design: To assess how BTLA and PD-1 signalling impact gene transcription induced by TCR/CD3 stimulation, we used a Jurkat-NFkB::eGFP reporter cell line together with a T-cell stimulator cell system that stimulates Jurkat T-cells via the TCR-CD3 complex. We stably co-expressed PD-1 or BTLA in our reporter cells and generated T-cell stimulator cells expressing the cognate ligands PD-L1 or HVEM. PD-1 or BTLA-expressing reporter cells were left unstimulated or co-cultured with control stimulator cells or stimulator cells expressing PDL-1 or HVEM. Following 4, 20 and 48 hours of co-culture, deep RNA-seq was used to distinguish and quantify changes in gene expression in the reporter and stimulator cells. As we co-cultured Jurkat cells with stimulator cells, the count matrix contains both human (ENSG) and mouse (ENSMUSG) genes. Further gene filtering needs to be applied to the count matrix if the scope of analysis is only limited to the Jurkat cells.