Short tandem repeats are important contributors to silencer elements in T cells.

The action of cis-regulatory elements with either activation or repression functions underpins the precise regulation of gene expression during normal development and cell differentiation. Gene activation by the combined activities of promoters and distal enhancers has been extensively studied in normal and pathological contexts. In sharp contrast, gene repression by cis-acting silencers, defined as genetic elements that negatively regulate gene transcription in a position-independent fashion, is less well understood. Here, we repurpose the STARR-seq approach as a novel high-throughput reporter strategy to quantitatively assess silencer activity in mammals. We assessed silencer activity from DNase hypersensitive I sites in a mouse T cell line. Identified silencers were associated with either repressive or active chromatin marks and enriched for binding motifs of known transcriptional repressors. CRISPR-mediated genomic deletions validated the repressive function of distinct silencers involved in the repression of non-T cell genes and genes regulated during T cell differentiation. Finally, we unravel an association of silencer activity with short tandem repeats, highlighting the role of repetitive elements in silencer activity. Our results provide a general strategy for genome-wide identification and characterization of silencer elements.

Cancer cells induce immune escape via glycocalyx changes controlled by the telomeric protein TRF2.

Myeloid-derived suppressor cells (MDSCs) are immature myeloid cells with strong immunosuppressive activity that promote tumor growth. In this study, we describe a mechanism by which cancer cells control MDSCs in human cancers by upregulating TRF2, a protein required for telomere stability. Specifically, we showed that the TRF2 upregulation in cancer cells has extratelomeric roles in activating the expression of a network of genes involved in the biosynthesis of heparan sulfate proteoglycan, leading to profound changes in glycocalyx length and stiffness, as revealed by atomic force microscopy. This TRF2-dependent regulation facilitated the recruitment of MDSCs, their activation via the TLR2/MyD88/IL-6/STAT3 pathway leading to the inhibition of natural killer recruitment and cytotoxicity, and ultimately tumor progression and metastasis. The clinical relevance of these findings is supported by our analysis of cancer cohorts, which showed a correlation between high TRF2 expression and MDSC infiltration, which was inversely correlated with overall patient survival.

Epromoters function as a hub to recruit key transcription factors required for the inflammatory response.

Gene expression is controlled by the involvement of gene-proximal (promoters) and distal (enhancers) regulatory elements. Our previous results demonstrated that a subset of gene promoters, termed Epromoters, work as bona fide enhancers and regulate distal gene expression. Here, we hypothesized that Epromoters play a key role in the coordination of rapid gene induction during the inflammatory response. Using a high-throughput reporter assay we explored the function of Epromoters in response to type I interferon. We find that clusters of IFNa-induced genes are frequently associated with Epromoters and that these regulatory elements preferentially recruit the STAT1/2 and IRF transcription factors and distally regulate the activation of interferon-response genes. Consistently, we identified and validated the involvement of Epromoter-containing clusters in the regulation of LPS-stimulated macrophages. Our findings suggest that Epromoters function as a local hub recruiting the key TFs required for coordinated regulation of gene clusters during the inflammatory response.

Integration of high-throughput reporter assays identify a critical enhancer of the Ikzf1 gene.

The Ikzf1 locus encodes the lymphoid specific transcription factor Ikaros, which plays an essential role in both T and B cell differentiation, while deregulation or mutation of IKZF1/Ikzf1 is involved in leukemia. Tissue-specific and cell identity genes are usually associated with clusters of enhancers, also called super-enhancers, which are believed to ensure proper regulation of gene expression throughout cell development and differentiation. Several potential regulatory regions have been identified in close proximity of Ikzf1, however, the full extent of the regulatory landscape of the Ikzf1 locus is not yet established. In this study, we combined epigenomics and transcription factor binding along with high-throughput enhancer assay and 4C-seq to prioritize an enhancer element located 120 kb upstream of the Ikzf1 gene. We found that deletion of the E120 enhancer resulted in a significant reduction of Ikzf1 mRNA. However, the epigenetic landscape and 3D topology of the locus were only slightly affected, highlighting the complexity of the regulatory landscape regulating the Ikzf1 locus.