TRANSCRIPTIONAL CONTROL OF Ca2+ SIGNALING IN T CELLS

Abstract

Antigen presentation to T cells results in their activation through T Cell Receptor (TCR) stimulation, resulting in sustained elevation of cytosolic Ca2+ concentration critical for T cell activation. Sustained Ca2+ signals are important for the activation of Nuclear Factor of Activated T cells (NFAT), which is a key regulator of T cell activation through its transcriptional control of genes in multiple process including cytokine production, proliferation and differentiation(Rao, Luo, & Hogan, 1997). Recently it was shown that Stromal Interaction Molecule 1 (STIM1) inhibits plasma membrane Ca2+/ATPase 4 (PMCA4) function during T cell activation resulting in sustained elevation of Ca2+ signals(Ritchie, Samakai, & Soboloff, 2012). This interaction requires upregulation of both STIM1 and PMCA4. In this thesis, I hypothesize that changes in Ca2+ signals arising from transcriptional changes of STIM1 and PMCA are important for the efficient activation of T cells. In the first part of this thesis, I assess the transcriptional regulation of STIM1 and PMCA4. My in vitro studies show that expression of both proteins is regulated by the EGR family members, EGR1 and EGR4. Additionally, transcriptional regulation of PMCA inhibition by EGR1 and EGR4 is required for efficient activation of T cells. Interestingly, whereas significant roles for EGR1, EGR2 and EGR3 in T cell development and function have been established, a role for EGR4 has not, hitherto been elucidated. In the second half of this thesis, using qPCR, I reveal that EGR4 expression is stimulated by TCR engagement in primary double positive, CD4 and CD8 positive murine T cells. Further, EGR4-null mice exhibit shifts in early thymic development, although this does not affect the relative number of double or single positive T cells in the thymus. Interestingly, EGR4-null primary T cells exhibit normal Ca2+ entry, but fail to exhibit activation-induced inhibition of Ca2+ clearance. Although not all subsets of EGR1 and EGR4 null primary T cells exhibited decreased STIM1 expression, significant defects in proliferation, migration and/or cytokine production were observed upon stimulation in all populations, albeit to different extents. These findings reveal a two-faceted role in which EGRs regulate T cell development and function through both Ca2+-dependent and independent methods. I believe that these findings have important implications towards the general understanding of transcriptional control of Ca2+ signaling, as well as having a possible impact in the quest to advance therapies targeting immunological disorders.