Pannexin-1 hemichannel–mediated ATP release together with P2X1 and P2X4 receptors regulate T-cell activation at the immune synapse

Abstract

Engagement of T cells with antigenpresenting cells requires T-cell receptor (TCR) stimulation at the immune synapse.We previously reported that TCR stimulation induces the release of cellular adenosine-5-triphosphate (ATP) that regulates T-cell activation. Here we tested the roles of pannexin-1 hemichannels, which have been implicated in ATP release, and of various P2X receptors, which serve as ATP-gated Ca 2؉ channels, in events that control T-cell activation. TCR stimulation results in the translocation of P2X1 and P2X4 receptors and pannexin-1 hemichannels to the immune synapse, while P2X7 receptors remain uniformly distributed on the cell surface. Removal of extracellular ATP or inhibition, mutation, or silencing of P2X1 and P2X4 receptors inhibits Ca 2؉ entry, nuclear factors of activated T cells (NFAT) activation, and induction of interleukin-2 synthesis. Inhibition of pannexin-1 hemichannels suppresses TCR-induced ATP release, Ca 2؉ entry, and T-cell activation. We conclude that pannexin-1 hemichannels and P2X1 and P2X4 receptors facilitate ATP release and autocrine feedback mechanisms that control Ca 2؉ entry and T-cell activation at the immune synapse. (Blood. 2010; 116(18):3475-3484) IntroductionT-cell activation requires a sustained elevation of intracellular Ca 2ϩ levels, which is accomplished by Ca 2ϩ entry through calcium release-activated calcium (CRAC) channels that are composed of stromal interaction molecule 1 (STIM1) and Orai1 proteins. [1][2][3] Both proteins translocate to the immune synapse upon T-cell activation, where they mediate localized influx of extracellular Ca 2ϩ . 4 Ca 2ϩ entry contributes to the activation of nuclear factors of activated T cells (NFATs) that induce interleukin-2 (IL-2) gene expression and subsequent signaling events that lead to T-cell proliferation. [5][6][7] Recent studies have shown that extracellular adenosine-5Ј-triphosphate (ATP) regulates T-cell activation. [8][9][10] T cells release ATP in a controlled manner, as do other leukocytes, thereby facilitating intercellular communication and autocrine feedback regulation of cell function. [8][9][10][11][12][13] Stimulation of T cells by T-cell receptor (TCR) ligation, mechanical stimulation, membrane deformation, or osmotic stress induces the release of cellular ATP. 9,10,[13][14][15][16] T cells express the gap junction hemichannels pannexin-1, which can mediate ATP release and T-cell activation. 8,10 T-cell activation has been shown to involve P2X receptor subtypes. 8,10 The 7 mammalian P2X receptor family members (P2X1-7) are ATPgated ion channels. 17,18 All these receptors, with the exception of P2X5, can facilitate entry of Ca 2ϩ in response to stimulation by extracellular ATP, [18][19][20] thus suggesting that P2X receptors regulate T-cell activation by mediating Ca 2ϩ entry.T-cell activation during antigen recognition requires the formation of an immune synapse between T cells and antigen-presenting cells. The immune synapse is a complex structure with a limited number of TCRs, implying that TCR stimu...

Mitochondria Regulate Neutrophil Activation by Generating ATP for Autocrine Purinergic Signaling

“…This suggests that spatiotemporal dynamics of mitochondrial translocation within the cell can regulate PMN functions. Indeed, similar observations were made in T cells where the accumulation of mitochondria and panx1 at the immune synapse results in the physical proximity of the sites of ATP production and of ATP release, minimizing rate-limiting diffusion and thus facilitating rapid local purinergic signaling at the immune synapse (25)(26)(27)(28)43). Thus, we conclude that mitochondrial ATP production and the location of mitochondria relative to panx1 contribute to purinergic signaling in stimulated PMNs.…”

“…Recent work by others has shown that different sets of P2 receptors regulate Ca 2ϩ signaling in other cell types (29). P2X receptors function as ATP-gated ion channels that can facilitate Ca 2ϩ influx, and in T cells, P2X1, P2X4, and P2X7 receptors regulate influx of extracellular Ca 2ϩ and cell activation in response to T cell stimulation (27,28). PMNs express several members of the P2X receptor family in addition to the predominant P2Y 2 receptor (5, 7).…”

Chemotherapeutic Drugs Induce ATP Release via Caspase-gated Pannexin-1 Channels and a Caspase/Pannexin-1-independent Mechanism

“…Recent studies in Jurkat cells and normal human T cells have indicated that the T cell receptor (TCR) activation triggers a very rapid but transient (within minutes) release of ATP via both Panx1 channel-mediated and exocytotic pathways (33,58,59). Both ATP release mechanisms were dependent on the increase in cytosolic [Ca 2ϩ ] elicited by TCR activation.…”

“…Moreover, most types of tumor cells per se express different cassettes of P2 purinergic or P1 adenosine receptors that can modulate cell growth, survival, and sensitivity to pro-apoptotic mediators. Indeed, the leukemic Jurkat cells used in this study express several ionotropic P2 receptor subtypes, including P2X1, P2X4, and P2X7, and these receptors act as autocrine modulators of TCR signaling pathways that regulate growth and cytokine production (58,59). It is interesting to speculate whether autocrine activation of these receptors during chemotherapeutic drug-induced Panx1 activation may also modulate apoptotic progression in Jurkat or other leukemic cells.…”

Function and Regulation of Pannexin 1 Channels in the Vascular Endothelium

“…Pannexin proteins are found in three different isoforms Panx channels have been suggested to act as paracrine release channels that are strongly implicated in the release of purine nucleotides from cells [30,154,162,165,227].…”