Investigating the Role of Bromodomain and Extraterminal (BET) Family Proteins in Human T Cell Function

Abstract

The human immune system consists of a myriad cells of both innate and adaptive origin that work in exquisite balance to initiate, propagate and resolve inflammatory responses to both internal and external stimuli. The central role of the T cell compartment in the orchestration, maintenance and regulation of appropriate immune responses is well known, as are the consequences of dysregulation resulting in a range of inflammatory and autoimmune diseases. Despite this, the epigenetic processes underpinning the phenotypic and functional differentiation of these cells remains a key area of interest in the field, with many questions as yet unanswered. This research investigated the role of BRD2, BRD3 and BRD4, collectively referred to as the bromodomain and extraterminal (BET) family of epigenetic proteins, which facilitate transcription via the recognition of and binding to chromatin modifications, with the hypothesis that these proteins would play a key role in the propagation of various phenotypic and effector functions in both CD4+ and CD8+ T cells. Given the importance of the BET family that has been attributed to the generation of both proliferative and inflammatory responses in other cell types as a consequence of small molecule inhibition using a range of chemically distinct BET inhibitors, we utilised the small molecule BET inhibitor, I-BET151 to identify an important role for the tandem bromodomain modules of BRD2, BRD3 and BRD4 in the activation, proliferation and effector function of both the human CD4+ and CD8+ T cell compartments following T cell receptor (TCR)- mediated activation in primary cells isolated from healthy donors. We confirmed previous literature findings in other disease models, that BET inhibition decreased the proliferative response to activation, as well as potently inhibiting a range of pro- inflammatory mediators including granzyme B, interferon gamma (IFN) and interleukin 17 (IL-17). Additionally, we were able to investigate the effects of BET inhibition at the transcriptional level, confirming that reduction in effector molecule production was not an artefact inherent to the anti-proliferative effects of BETi, an observation which appeared to be driven at least to some extent by the inhibition of the transcription factor, MYC. Furthermore, we identified nuanced phenotypic effects resulting from the selective inhibition of either the first (bromodomain 1, BD1) or second (bromodomain 2, BD2) of the tandem bromodomain modules of the BET family proteins, reporting herein for the first time, the differential effects of inhibiting the function of these separate modules in the CD8+ T cell compartment. Taken together, these data highlight an important role for the function of BET family proteins in underpinning the conveyance of T cell- mediated effector function in humans and provide new insights into the differential roles played by the individual bromodomains within these proteins in the context of TCR- mediated activation. These findings suggest that both pan- and domain selective- BET inhibition may offer new avenues of therapeutic value in the field of inflammatory and autoimmune disease treatment.