AbstractAfter transcriptional activation on encountering a stimulus, memory T cells can “remember” their transcription profile by priming genes with active epigenetic signatures to remain in a non-transcribing state throughout subsequent cell divisions. This is coupled with the ability to induce more efficient and robust transcription upon antigenic re-exposure, a phenomenon known as transcriptional memory (TM). The TM response is facilitated by permissive chromatin structures. However, the molecular details of how the permissive epigenetic landscape incorporates incoming stimulatory signals for transcriptional priming in human memory T cells are poorly defined.
To better understand the process of T cell response priming, formaldehyde-assisted isolation of regulatory elements-sequencing (FAIRE-seq) was used to globally profile chromatin accessibility in a previously described human Jurkat T cell transcriptional memory (JTM) model representing four distinct transcriptional states: (1) non-stimulated cells, (2) stimulated cells, (3) stimulus withdrawal cells with a resting memory state, and (4) re-stimulated cells. Initial stimulation altered the chromatin accessibility landscape, which largely persisted throughout subsequent cell divisions, and secondary activation induced secondary-specific enhanced accessibility at previously restricted regions. Increased accessibility occurred largely in distal regulatory regions and was characterized by increased enrichment of the H3K27ac and H3K4me active enhancer markers. Chromosome conformation capture (3C) analysis showed that the enhancer-promoter chromatin interaction existed prior to initial activation in non-stimulated T cells. These pre-established chromatin loops may contribute to the rapid transcription priming response of “poised” genes upon stimulation.
Protein kinase C-theta (PKC-θ) plays a key role in effective human T cell activation and requires T cell receptor engagement with co-stimulatory signalling. In vitro T cell stimulation with phorbol 12-myristate 13-acetate (PMA) and calcium ionophore (CaI) mimics human T cell activation and induces immune responsive gene transcription via PMA-activated PKC-θ signalling. Here, the enhanced TM response was associated with the strength of initial PKC-θ signalling, and PKC-sensitive regions showed higher chromatin accessibility upon secondary stimulation. Moreover, blocking the nuclear translocation of PKC-θ using a cytoplasm-restricted PKC-θ mutant highlighted the importance of nuclear PKC-θ in maintaining the permissive chromatin state of TM genes by H2B serine 32 phosphorylation. Overall, transcriptional responses are primed via the chromatin accessibility landscape in memory T cells.
|Date of Award||2018|
|Supervisor||Sudha Rao (Supervisor), Ian Ramshaw (Supervisor) & Kris Hardy (Supervisor)|