Nature Immunology
- 7, 1132 - 1134 (2006)
doi:10.1038/ni1106-1132
Linking diacylglycerol kinase to T cell anergyDaniel L MuellerDaniel L. Mueller is in the Department of Medicine and Center for Immunology, University of Minnesota Medical School, Minneapolis, Minnesota 55455 USA. muell002@umn.edu Understanding of the signaling networks regulating T cell anergy remains incomplete. Two reports now demonstrate that alterations in diacylglycerol metabolism regulate the adoption of an anergic versus an activated T cell fate.Just over 20 years ago, it was reported that human cloned T cells enter a state of unresponsiveness when allowed to present antigen to each other1. Shortly thereafter, an analogous form of tolerance was noted in vitro and in vivo in mouse T cells exposed to antigen in the presence of 'costimulation-poor', chemically modified antigen-presenting cells2 or purified major histocompatibility complex class II molecules3. In both experimental systems, 'anergic' T cells were incapable of proliferating after rechallenge with antigen, at least in part because of their inability to produce interleukin 2 (IL-2). Coincidentally, at about the same time, involvement of inositol phospholipid hydrolysis and the accumulation of diacylglycerol as a second messenger for receptor-mediated protein kinase C activation was identified4. In this issue Nature Immunology, two reports5,
6 now provide insight into the biochemical mechanisms underlying the development and maintenance of the anergic state and identify a central function for diacylglycerol in this regulation of immunological tolerance.
Gajewski and colleagues built on the observation that the small GTP-binding protein Ras becomes uncoupled from the T cell receptor after the induction of T cell anergy7. That defect in Ras signaling seems to be important, as anergic T cells fail to properly activate 'downstream' mitogen-activated protein kinases and have defective transactivation of Il2 dependent on the transcription factor AP-1 (ref. 8). Consistent with that idea, these authors now show that the ectopic expression of a constitutively active, mutant form of Ras in anergic T cells overcomes many of those functional defects and restores IL-2 secretion and proliferation in response to antigen rechallenge5. Notably, diacylglycerol-dependent recruitment of the Ras-activating guanine nucleotide–exchange factor RasGRP1 to the immunological synapse is defective in anergic T cells, and anergic T cells contain excessive diacylglycerol kinase- (DGK-), which phosphorylates diacylglycerol to form phosphatidic acid. The addition of a DGK--specific chemical inhibitor during rechallenge with agonistic antibodies specific for CD3 and CD28 substantially improved IL-2 production, suggesting that an abundance of DGK- accounts at least in part for the functional defects noted in anergic T cells.
Koretzky and colleagues have used a genetic knockout approach to examine the involvement of diacylglycerol metabolism in the regulation of T cell anergy6. T cells lacking either DGK- or DGK- develop normally and populate secondary lymphoid organs. However, stimulation of Dgka-/- T cells elicited modest increases in Ras GTP-binding and 'downstream' kinase activation. Dgka-/- T cells produced more IL-2 and proliferated more in response to T cell receptor ligation, but resembled wild-type T cells in their response to a combination of calcium ionophore and a diacylglycerol analog that is insensitive to diacylglycerol kinases. Perhaps most notably, Dgka-/- T cells were relatively insensitive to the tolerance-inducing effects of chronic in vivo stimulation with the staphylococcal enterotoxin B 'superantigen'. Such resistance to anergy induction was also noted in vitro in T cell receptor–stimulated DGK--deficient T cells treated with a pharmacological inhibitor of DGK-.
These two studies offer a cogent model of immune tolerance that relies on the active suppression of an essential stimulatory pathway in anergic T cells (Fig. 1). As a consequence of anergy induction, DGK molecules are more readily available to associate with RasGRP1-Ras complexes at the immunological synapse, where they promote the conversion of nearby diacylglycerol molecules to phosphatidic acid. In the presence of lower local concentrations of diacylglycerol, RasGRP1 fails to effectively catalyze the loading of Ras with GTP, and recruitment of Raf to the RasGRP1-Ras complex does not occur. The end result is suboptimal activation of 'downstream' mitogen-activated protein kinases as well as ineffective IL-2 production and proliferation. Some precedent for such a model exists, as CD25+ regulatory T cells, which are important in maintaining self-tolerance, also have a block in autocrine proliferation, and their proliferative defect can be at least partially overcome by pharmacological inhibition of DGK activity9.
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How well does this model fit with existing data on various signaling alterations in T cells either during or after the induction of anergy? In some situations, diacylglycerol can increase during T cell activation independently of the delivery of costimulatory signals or the induction of anergy10. Furthermore, anergy can be induced even after the retroviral transduction of T cells with a constitutively active Ras construct11. Finally, clonal anergy develops after antigen stimulation even in the presence of CD28 costimulatory signals, if cell cycle progression is interrupted by inhibition of the mammalian target of rapamycin12. Thus, it seems unlikely that clonal anergy can occur only in the absence of diacylglycerol accumulation and/or Ras activation.
However, it is conceivable that alterations in diacylglycerol metabolism underlie many of the signaling and phenotypic differences noted before in anergic T cells. Diacylglycerol binds with high affinity to many proteins containing a cysteine-rich 1 domain, including RasGRP1 and DGK, as well as protein kinase C- and protein kinase D. Both protein kinase C- and protein kinase D are recruited to and are activated at the immunological synapse during antigen recognition in response to the hydrolysis of phosphatidylinositol-4,5-bisphosphate and the accumulation of diacylglycerol13. Therefore, enhanced DGK activity in anergic cells may limit the capacity of protein kinase C- to trigger 'downstream' gene expression mediated by transcription factor NF- B and may also interfere with the normal phosphorylation and nuclear export of histone deacetylases by activated protein kinase D. Both inhibitory events have the potential to act in synergy with defects in signaling 'downstream' of Ras to contribute to the maintenance of T cell anergy.
Enhanced DGK activity in the setting of the hydrolysis of phosphatidylinositol-4,5-bisphosphate in the immunological synapse could also be expected to lead to overproduction of polyunsaturated fatty acid–containing phosphatidic acid molecules that can act as second messengers elsewhere in the T cell. Excess phosphatidic acid–derived polyunsaturated fatty acids may be incorporated into the nuclear inositol phospholipid pool and may modulate histone acetylation and cell cycle progression through direct interaction with nuclear proteins containing plant homeodomain fingers14. Such long-range effects of altered diacylglycerol metabolism on chromatin accessibility might even contribute to the unique pattern of gene expression already noted after the induction of T cell anergy15. Alternatively, anergy may ultimately represent a differentiation state that leads to loss of autocrine growth factor production and proliferation as a consequence of the new expression of many distinct counter-regulatory molecules (such as Cbl-b, Itch and Grail), with DGK being just one more factor needed to prevent anergy reversal and maintain the tolerant state. Regardless of which is true, this identification of DGK as an important mediator of clonal anergy now provides the basis for new therapeutic approaches to the treatment of clinical autoimmunity that take advantage of diacylglycerol antagonism in T cells. Likewise, vaccines designed to promote diacylglycerol formation and limit the accumulation of DGK activity will have the greatest chance of eliciting a durable and protective response by antigen-specific T cells.
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