STAT5-mediated chromatin interactions in superenhancers activate IL-2 highly inducible genes: Functional dissection of the Il2ra gene locus

Abstract

Cytokines critically control immune responses, but how regulatory programs are altered to allow T cells to differentially respond to distinct cytokine stimuli remains poorly understood. Here, we have globally analyzed enhancer elements bound by IL-2-activated STAT5 and IL-21-activated STAT3 in T cells and identified as the top-ranked gene regulated by an IL-2-activated STAT5-bound superenhancer and one of the top genes regulated by STAT3-bound superenhancers. Moreover, we found that STAT5 binding was rapidly superenriched at genes highly induced by IL-2 and that IL-2-activated STAT5 binding induces new and augmented chromatin interactions within superenhancer-containing genes. Based on chromatin interaction analysis by paired-end tag (ChIA-PET) sequencing data, we used CRISPR-Cas9 gene editing to target three of the STAT5 binding sites within the superenhancer in mice. Each mutation decreased STAT5 binding and altered IL-2-induced gene expression, revealing that individual elements within the superenhancer were not functionally redundant and that all were required for normal gene expression. Thus, we demonstrate cooperative utilization of superenhancer elements to optimize gene expression and show that STAT5 mediates IL-2-induced chromatin looping at superenhancers to preferentially regulate highly inducible genes, thereby providing new insights into the mechanisms underlying cytokine-dependent superenhancer function.

Targeting STAT3 and STAT5 in Tumor-Associated Immune Cells to Improve Immunotherapy

“…Transcriptomic analyses of STAT5ca-expressing CD8 T cells highlighted a role for STAT5ca in the stabilization of a broad Tc-1 gene expression program initiated by TCR stimulation [60] (see Table 2, Figure 2). This observation is in agreement with the reported chromatin interactions of STAT5 in super-enhancers to activate IL-2 highly inducible genes [71]. .…”

“…Intriguingly, IL-21 was shown to promote in vitro proliferation of CD8 T naïve and memory cells in synergy with STAT5-inducing cytokines IL-7/IL-15 [90]. Indeed, Leonard's group reported that IL-2 vs. IL-21 cytokine stimulation of pre-activated T cells induces STAT5 and STAT3 binding, respectively, to incompletely overlapping super enhancers [71]. Understanding the fine-tuning of cytokine-modulated STAT3/5 activities will require further effort as reviewed in [46]: This tight regulation relies on the T cell context, including cell lineage, differentiation state, and cytokine milieu, triggering synergistic/antagonistic STAT-mediated transcriptional regulation.…”

Progressing super-enhancer landscape during mammary differentiation controls tissue-specific gene regulation

“…STAT5 is not only central to hematopoietic lineages and mammary luminal cells that depend on cytokines ( 44 , 45 ), but also modulates gene expression in other cell types. The precise and cell-specific roles of STAT5-based enhancers might depend on context, possibly their relative position within super-enhancers, relative distance to promoters or accessibility in permissive chromatin ( 10 , 39 , 46 , 47 ). Chromatin accessibility is considered as a key to transcription factor binding and activating gene expression ( 48 ).…”

Chromatin Accessibility and Interactions in the Transcriptional Regulation of T Cells

“…This differential STAT activation leads to differential gene expression by these cytokines. It is established that STAT proteins are critical components of cytokine-activated enhancers, but recently their roles related to super-enhancers ( 70 ) and their abilities to fine-tune gene expression ( 71 ) have been elucidated, with, for example, greater IL-2-inducibility of genes with STAT5-based super-enhancers, as compared to STAT5-based typical enhancers ( 71 ). As opposed to typical enhancers, where factor binding occurs in more limited regions, super-enhancers (also known as stretched or clustered enhancers) ( 70 , 72 , 73 ) represent groups of putative enhancers in close genomic proximity that span broader regions (Figure 2 ), are densely bound by transcriptional coactivators, and usually are associated with high levels of the active chromatin mark histone H3 lysine 27 acetylation (H3K27Ac).…”