Integration of Kinase and Calcium Signaling at the Level of Chromatin Underlies Inducible Gene Activation in T Cells.

Ruth Brignall,Lorraine Schmidt,Nigel P Dyer,James Bagnall,Sarah L Bevington,Angela O Pisco,Pierre Cauchy,Peter N Cockerill,William Rowe,Pawel Paszek,Mark A Travis,Bethany Gorman,Christopher Boddington,Sascha Ott,Dean A Jackson,Hazel England,Kevin Rich

doi:10.4049/jimmunol.1602033

Abstract

TCR signaling pathways cooperate to activate the inducible transcription factors NF-κB, NFAT, and AP-1. In this study, using the calcium ionophore ionomycin and/or PMA on Jurkat T cells, we show that the gene expression program associated with activation of TCR signaling is closely related to specific chromatin landscapes. We find that calcium and kinase signaling cooperate to induce chromatin remodeling at ∼2100 chromatin regions, which demonstrate enriched binding motifs for inducible factors and correlate with target gene expression. We found that these regions typically function as inducible enhancers. Many of these elements contain composite NFAT/AP-1 sites, which typically support cooperative binding, thus further reinforcing the need for cooperation between calcium and kinase signaling in the activation of genes in T cells. In contrast, treatment with PMA or ionomycin alone induces chromatin remodeling at far fewer regions (∼600 and ∼350, respectively), which mostly represent a subset of those induced by costimulation. This suggests that the integration of TCR signaling largely occurs at the level of chromatin, which we propose plays a crucial role in regulating T cell activation.

Highlights

In this study, using the calcium ionophore ionomycin and/or PMA on Jurkat T cells, we show that the gene expression program associated with activation of TCR signaling is closely related to specific chromatin landscapes
In agreement with previous analyses [70,71,72], stimulation with PMA alone resulted in NF-kB pathway activation whereas stimulation with ionomycin alone led to NFAT activation (Fig. 1C, 1D)
To investigate transcriptional responses to PMA and ionomycin, we employed Jurkat T cells transfected with luciferase reporter gene lentiviruses, driven by either the full-length NFAT/AP-1– dependent IL-2 promoter (IL-2–Luc) or an array of five NF-kB sites (5kB-Luc)

Summary

Introduction

In this study, using the calcium ionophore ionomycin and/or PMA on Jurkat T cells, we show that the gene expression program associated with activation of TCR signaling is closely related to specific chromatin landscapes. The binding motif for NF-kB was detected in 12% of these inducible DHSs. The activation of gene expression [27, 28] and the establishment of immunological memory [1, 29,30,31,32] in T cells are controlled by TFs that recruit chromatin modifiers and remodelers to transcriptional enhancers and promoters.

Results

Conclusion