A signal-switch hypothesis for cross-regulation of cytokine and TLR signalling pathways

Abstract

The importance of immunoreceptor tyrosine-based activation motif (ITAM)-coupled receptors in modulating signalling pathways downstream of other types of receptor is well established, but the mechanisms underlying this modulation are not known. Recent data suggest that calcium-dependent signalling downstream of ITAM-coupled receptors regulates the amplitude and functional outcomes of cytokine and TLR signalling. In this Opinion article, I describe a model whereby the intensity of ITAM-dependent signalling and the balance of calcium signals relative to other ITAM-mediated signalling pathways determines whether cellular responses to cytokines and TLR ligands are increased or inhibited. This model describes mechanisms that explain how ITAM-coupled receptors regulate heterologous signalling pathways.

Figure 1. Canonical ITAM-mediated signalling

Crosslinking of immunoreceptor tyrosine-based activation motif (ITAM)-associated receptors (such as triggering receptor expressed on myeloid cells 2 (TREM2), Fc receptors (FcRs) or β2-integrin) leads to SRC family kinase-dependent phosphorylation of tyrosines in the ITAM motif in the cytoplasmic domain of ITAM-containing adaptor molecules, followed by recruitment and activation of a ζ-chain-associated protein kinase of 70 kDa (ZAP70) or spleen tyrosine kinase (SYK). In myeloid cells, FcRγ and DNAX activation protein 12 (DAP12) are the major ITAM-containing adaptors and spleen tyrosine kinase (SYK) is the major recruited kinase. SYK initiates a signalling cascade that leads to activation of Bruton’s tyrosine kinase (BTK), the formation of multiprotein signalling complexes coordinated by the scaffolding adaptors B-cell linker (BLNK) and SH2-domain-containing leukocyte protein of 76 kDa (SLP76) and activation of phospholipase Cγ (PLCγ). PLCγ activates key downstream effector pathways through generation of the second messengers diacylglycerol (DAG) and inositol-1,4,5-trisphosphate (InsP₃) that activate the depicted kinase cascades, such as RAS guanyl-releasing protein (RasGRP), mitogen-activated protein kinases (MAPKs), calmodulin-dependent kinase (CaMK) and protein tyrosine kinase 2 (PYK2), and downstream transcription factors such as nuclear factor-κB (NF-κB), activator protein 1 (AP-1), cAMP-responsive-element-binding protein (CREB), myocyte enhancer factor 2 (MEF2) and nuclear factor of activated T cells (NFAT).

Figure 2. Tonic calcium signalling by low-avidity ligation of ITAM-coupled receptors modulates cytokine and TLR signalling

Low-avidity ligands, such as monomeric immunoglobulin G (IgG) or IgA that bind Fc receptors (FcRs) or endogenous triggering receptor expressed on myeloid cells 2 (TREM2) ligands (including semaphorin 6D), are sufficient to induce tonic calcium signalling that leads to basal activity of calmodulin-dependent kinase (CaMK), protein tyrosine kinase 2 (PYK2) and calcineurin. PYK2 interacts with Janus kinases (JAKs) and enhances interferon (IFN)-induced JAK–STAT (signal transducer and activator of transcription) signalling, whereas calcineurin interacts with Toll-like receptors (TLRs) and the adaptor molecule MyD88 (myeloid differentiation primary-response gene 88) and attenuates TLR signalling. Calcineurin-mediated activation of nuclear factor of activated T cells (NFAT) and downstream target genes that include phosphatases, E3 ubiquitin ligases and transcriptional repressors may contribute to inhibition. SH2-domain-containing protein tyrosine phosphatase 1 (SHP1) is recruited to partially and/or weakly phosphorylated immunoreceptor tyrosine-based activation motifs (ITAMs) and can cross-inhibit TLRs. Low-avidity ligands do not substantially activate nuclear factor-κB (NF-κB) or mitogen-activated protein kinase (MAPK) pathways.

Figure 3. High-intensity signalling after high-avidity ligation of ITAM-coupled receptors inhibits cytokine signalling but synergizes with TLR signalling

High-avidity ligands, such as immune complexes that bind Fc receptors (FcRs), strongly and acutely activate signalling pathways leading to the activation of protein kinase C (PKC), mitogen-activated protein kinases (MAPKs), nuclear factor-κB (NF-κB) and reactive oxygen species (ROIs). MAPKs, NF-κB and ROIs enhance and synergize with Toll-like receptor (TLR) signals in the inflammatory activation of macrophages. PKC has a key role in the inhibition of cytokine JAK–STAT (Janus kinase–signal transducer and activator of transcription) signalling by inducing recruitment of SH2-domain-containing protein tyrosine phosphatase 1 (SHP2) to the type I interferon (IFN) receptor IFNAR, and by inducing internalization of the interleukin-10 receptor (IL-10R). In T cells, activation of PKC downstream of ζ-chain-associated protein kinase of 70 kDa (ZAP70) (homologous to the myeloid-expressed protein SYK) inhibits cytokine signalling through an extracellular-signal-regulated kinase (ERK)-dependent mechanism; in parallel, acute activation of calcineurin contributes to inhibition of IL-4R signalling. Activation of p38 MAPK in macrophages by immunoreceptor tyrosine-based activation motif (ITAM)-coupled receptors such as TLRs, IL-1R and tumour-necrosis factor receptor (TNFR) strongly inhibits signalling by cytokines that use the gp130 signalling receptor subunit such as IL-6 and IL-27. p38-MAPK-dependent inhibition of gp130 signalling is mediated by induced recruitment of SHP2 and receptor internalization and indirectly through induction of the signalling inhibitor suppressor of cytokine signalling 3 (SOCS3).TCR, T-cell receptor

Figure 4. Opposing effects of low- and high-avidity ligation of ITAM-coupled receptors are mediated by differential engagement of signalling pathways

(a) Macrophages sense the cell and tissue environment through interactions of immunoreceptor tyrosine-based activation motif (ITAM)-coupled receptors with low-avidity ligands. Examples include ligation of triggering receptor expressed on myeloid cells 2 (TREM2) by its tissue-expressed ligand semaphorin 6D and as yet unknown macrophage-expressed ligands and of Fc receptors (FcRs) by monomeric immunoglobulin G (IgG) in the serum. This low-avidity signal is tranduced through spleen tyrosine kinase (SYK), phospholipase Cγ (PLCγ) and inositiol-1,4,5-triphosphate (InsP₃) to modulate calcium concentrations and thus regulate the basal activity of calmodulin-dependent kinase (CaMK), protein tyrosine kinase 2 (PYK2) and calcineurin that modulate Toll-like receptor (TLR) and cytokine-mediated responses to make them appropriate for the cell environment. (b) Dramatic and acute changes in the environment (for example, formation of immune complexes that bind FcRs during infection) can lead to high-avidity ligation of ITAM-coupled receptors and strong activation of protein kinase C (PKC), mitogen-activated protein kinases (MAPKs) and nuclear factor-κB (NF-kB). The switch to a PKC-, MAPK- and NF-κB-dominant signal promotes synergy with TLRs and inhibits cytokine signalling.