TCR, BCR, Soluble Tyrosine Kinases and NFAT

Initially, the discovery of detergent-resistant fractions in membranes obtained from epithelial cells28 led to the postulation of membrane regions enriched in glycosphingolipids and cholesterol. These were also referred to as detergentresistant (or detergent-insoluble) membrane domains, or more simply as lipid rafts. They are thought to be phase-separated regions, which, because of the saturated nature of the acyl chains of the component sphingolipids and the presence of cholesterol, possess a higher order of rigidity than the remainder of the membrane. Direct visualization of lipid microdomains in a smooth muscle cell by ‘single molecule’ fluorescence microscopy indicates dimensions ranging between 0.2 and 2  m at ambient temperatures, but at 37 °C they may be smaller.29 Both their size and density are likely to vary with cell type.

In the context of this chapter, signalling molecules that appear to be confined to lipid rafts include CD4/CD8 in T cells and the dual acylated (N-myristoylated and palmitoylated) non-receptor tyrosine kinases of the Src family. The nonacylated Src kinase Lck is also raft-associated, presumably through association with CD4/CD8. Other important signalling components located in rafts are LAT and CARMA1 (see above). Membrane proteins such as the transferrin receptor and the tyrosine phosphatase CD45 are generally absent. The primary receptors – the TCR, the BCR, and the IgE-R – were thought to exist outside rafts until cross-linked during activation, but it now appears that a proportion of these receptors is associated with rafts under resting conditions and that this increases following stimulation. Exactly how lipid rafts function as platforms for the assembly of signalling complexes remains unclear.

Signalling through interferon receptors

Classification of the interferons (IFN), cytokines originally recognized for their antiviral properties,30 has been difficult. The main classes and subtypes, and their receptors are listed in Table 17.1 and illustrated in Figure 17.8.

The interferons are classified as types I, II, and III. The type I subfamily contains 8 members that all bind to the same receptor heterodimer composed of IFNAR-1 and -2. The type II interferon, represented uniquely by IFN- , interacts with IFNGR-1 and -2. Type III interferons comprise three variants of IFN-l, each binding to the interleukin 28Ra/10R2 receptor dimer.

Hours to days before the adaptive immune response gets under way to combat viral infection, IFN- , - , and - are already at work.33 Mice that lack the receptors for IFN- , - , and - exhibit increased susceptibility to viral infection.34 The interferons induce the expression of protein kinase-R (dsRNA-dependent

protein kinase), 2 –5 oligoadenylate synthetases (OAS) and methyl transferases. These (respectively) bring about (either directly or indirectly) arrest of protein synthesis, degradation of single-stranded RNA, and inhibition of transcription, thus hindering the propagation of the virus. In some cases, interferons cause

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Table 17.1  Classification of interferons and their receptors

Adapted from Takaoka and Yanai.31

Fig 17.8  Interferons and their receptors.

The figure illustrates the organization of IFNAR1 and IFNAR2 bound to IFN- . (1n6v32).

Adapted from Chill et al.32

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Fig 17.9  Activation of STATS by the interferon receptor.

Binding of IFNcauses dimerization of the IFNreceptor (IFNARs 1 and 2) (1). This brings together the associated cytosolic kinases TYK2 and JAK1 which phosphorylate each other (2) and then IFNAR1 (3). STAT2, which in its latent form is bound to IFNAR2, associates with the tyrosine phosphorylated IFNAR1 through its SH2 domain and in so doing it too becomes phosphorylated (4). The phosphorylated STAT2 catalyses the phosphorylation of STAT1 and these detach from the receptor complex. The dimer is accompanied (5) by an interferon regulatory factor IRF-9. The ternary complex, ISGF3, enters the nucleus (6) and binds to DNA at the interferon stimulated response element (ISRE). The complex induces expression of genes such as interferon regulatory factors (IRF), ssRNA kinase, methyltransferases (Mx), and the chemokine IP-10.

cell death through apoptosis.33 IFN- , - , and - also play essential roles in immune modulation. They promote maturation of dendritic cells by increasing surface expression of costimulatory molecules and MHC class I and II. As a result, the ability of dendritic cells to stimulate the clonal expansion of effector T cells (e.g. Th1) is enhanced. In certain cases they may induce tolerance.35

Besides their roles in viral defence, IFNand - are important in the control of differentiation and growth. Generated in the bone marrow, they affect the differentiation of haematopoietic B cells, T cells, osteoclasts, and myeloid dendritic cells.36 IFN- 2 is used in adjuvant therapy as an anticancer drug. In particular, it has been applied in the treatment of ‘hairy cell leukaemia’ (a chronic B cell leukaemia37), chronic myelogenous leukaemia,38 and to treat some metastasizing cancers, such as renal carcinoma and Kaposi’s sarcoma, which affects AIDS sufferers.

Interferonreceptor and STAT proteins

The interferon- (type 1) receptor is made up of two subunits, IFNAR1 and IFNΑR2, coupled to the FERM domain of TYK2 and JAK1 respectively39 (Figure 17.9). Both are members of the Janus kinase family. IFNAR2 also binds

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