Calcium channels in lymphocytes

On B and T lymphocytes, ligation of the antigen receptor (AgR) induces a biphasic Ca2+ response. In the initial phase there is a large elevation in the intracellular Ca2+ concentration as a consequence of Ca2+ release from intracellular stores. This is followed by a lower, but prolonged elevation that is dependent on extracellular Ca2+. 1,2 This simple description belies the complexity of the response. The initial phase may involve as many as three different intracellular Ca2+ channels, while the second phase depends not only on plasma membrane Ca2+ channels, but also on at least two different intracellular channels. The complexity of the signal, and the many opportunities for regulation of individual components of the signalling mechanism, lead to a tremendous flexibility in outcome, ranging from single, brief elevated Ca2+ transients, through a range of oscillatory responses, each of which can be decoded by the cell into a differing outcome.2 In this review we concentrate on the Ca2+ channels involved in the AgR-mediated Ca2+ signal, but we briefly discuss other Ca2+ channels present in lymphocytes. Figure 1 shows two possible schemes for the involvement of Ca2+ channels in TCR signalling, and Fig. 2 shows possible roles for Ca2+ channels in B cells. Figure 1 A possible scheme for the involvement of Ca2+ channels in TCR signalling (a) depicts the simplest possible scheme for the role of Ca2+ channels in TCR-induced Ca2+ signalling. TCR-induced Ins(1,4,5)P3 production causes Ca2+ release from intracellular . Figure 2 Possible roles for Ca2+ channels in B cells. The BCR-induced Ca2+ signal involves the production of Ins(1,4,5)P3 and the release of Ca2+ from intracellular stores gated by InsP3Rs and RyR1. This is followed by an influx of Ca2+ through an unidentified . INTRACELLULAR Ca2+ CHANNELS A plethora of studies of AgR signalling have highlighted the role of inositol trisphosphate [Ins(1,4,5)P3]-mediated release of Ca2+ from internal stores (reviewed in refs 1–3). However, it is becoming apparent that there is more to the regulated release of intracellular Ca2+ in lymphocytes than inositol trisphosphate receptors (InsP3Rs). Recent studies are beginning to unravel roles for ryanodine receptors (RyRs) and the newly described and little understood NAADP receptor. Inositol trisphosphate receptors Three types of InsP3R are known, and they vary in their sensitivities to Ins(1,4,5)P3 and in the properties of their activation by Ca2+. InsP3Rs must bind Ins(1,4,5)P3 for Ca2+ release to occur. The response of the InsP3R can be regulated by phosphorylation, by various accessory proteins and by ATP, but by far the most important regulator is Ca2+. The exact mechanism is disputed4–6 but it is apparent that the differing sensitivities of the InsP3R isoforms to regulation by Ca2+ allow cells to fine-tune the temporal and spatial aspects of the Ca2+ signal.5 Much recent work has been directed towards determining the roles of the various isoforms. B and T cells express all three types of InsP3R to varying degrees depending on their stage of differentiation.7–11 It is not clear why lymphocytes simultaneously express all three isoforms, particularly since Sugawara et al.8 clearly demonstrated the redundancy of InsP3R expression. In a series of knockouts in DT40 B lymphocytes the BCR-induced Ca2+ signal could not be ablated until all three InsP3R isoforms were simultaneously knocked out.8 InsP3R3 was the first isoform to have a specific role ascribed to it. In both T and B cells, InsP3R3 is up-regulated in cells undergoing apoptosis.10 Inhibition of InsP3R3 expression using antisense RNA prevents TCR-induced apoptosis.10 However, Sugawara et al.8 provided convincing evidence against a specific role for InsP3R3 in apoptosis when they showed that knockout of any two isoforms in DT40 cells inhibited BCR-induced apoptosis. InsP3R3 can be expressed on the external surface of the plasma membrane of T and B cells and in T cells it can cocap with the TCR.9,10 The validity of this observation has been questioned, but support was provided by Tanimura et al.12 who demonstrated InsP3R3 expression on the external surface of Jurkat T cells. They showed that this pattern of expression was not limited to InsP3R3, and in fact the predominant isoforms in the plasma membrane were InsP3R1 and InsP3R2.12 The role of plasma membrane InsP3Rs is unknown, but Putney13 has suggested that, in some cell types, InsP3R3 may be expressed as an integral plasma membrane protein and function as all or part of a store-operated Ca2+ channel. However, the properties of InsP3R, in particular its Ca2+ selectivity profile, do not accord with those of known store-operated Ca2+ channels and the prevailing evidence strongly indicates that these channels are formed by distinct molecules from InsP3Rs (discussed below). In an attempt to explore the role of InsP3R1, Jayaraman et al.11 used antisense RNA to show that TCR-induced Ca2+ signals are exclusively transduced through this subtype. However, this is a controversial finding since the construct they used could cross-react with the type 2 and type 3 receptors. Hirota et al.14 subsequently demonstrated that T lymphocytes could develop normally in InsP3R1 knockout mice and showed normal Ca2+ fluxes in response to anti-CD3 stimulation, contradicting the claim that InsP3R1 was absolutely required for TCR signalling. In the most convincing study to ascribe roles for the various isotypes, Miyakawa et al.15 showed that InsP3R3 was involved in the generation of monophasic single Ca2+ transients following BCR ligation, whereas InsP3R1 and InsP3R2 were involved in the generation of Ca2+ oscillations with differing frequencies. The knockouts studied by Sugawara et al.8 suggest that inhibition of downstream events may be achieved simply by reducing the overall levels of InsP3Rs, rather than the specific levels of one particular isotype. This raises the possibility that cells do not express homotetramers of InsP3Rs; rather they may express heterotetramers of varying amounts of each isoform, allowing the cell to express a graded array of hybrid receptors with a wide variety of subtly different properties made up of combinations of the properties of the ‘pure’ homotetramers. Clearly much remains to be done to unravel the role of InsP3Rs in AgR signalling.