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SOCS3 binds specific receptor–JAK complexes to control cytokine signaling by direct kinase inhibition

Abstract

The inhibitory protein SOCS3 plays a key part in the immune and hematopoietic systems by regulating signaling induced by specific cytokines. SOCS3 functions by inhibiting the catalytic activity of Janus kinases (JAKs) that initiate signaling within the cell. We determined the crystal structure of a ternary complex between mouse SOCS3, JAK2 (kinase domain) and a fragment of the interleukin-6 receptor β-chain. The structure shows that SOCS3 binds JAK2 and receptor simultaneously, using two opposing surfaces. While the phosphotyrosine-binding groove on the SOCS3 SH2 domain is occupied by receptor, JAK2 binds in a phosphoindependent manner to a noncanonical surface. The kinase-inhibitory region of SOCS3 occludes the substrate-binding groove on JAK2, and biochemical studies show that it blocks substrate association. These studies reveal that SOCS3 targets specific JAK–cytokine receptor pairs and explains the mechanism and specificity of SOCS action.

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Figure 1: The structure of a JAK–SOCS–gp130 complex and the two interfaces.
Figure 2: The SOCS3-JAK2 interaction.
Figure 3: The JAK2-binding site is composed of the KIR, ESS and SH2 domain and is conserved between SOCS3 and SOCS1.
Figure 4: The KIR is required for JAK binding.
Figure 5: SOCS3 inhibits JAK2 by blocking substrate binding.
Figure 6: Residues upstream of the SOCS3 KIR act as a pseudosubstrate.
Figure 7: Comparison of three kinase inhibitors: SOCS3, PAK1IS and Grb14.

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Acknowledgements

We thank P. Colman, P. Czabotar and M. Lawrence for advice and I. Segel for helpful discussions. This work was supported in part by the National Health and Medical Research Council of Australia (NHMRC), program grants 461219 and 1016647 (N.A.N.) and project grant 1011804 (J.J.B.), the US National Institutes of Health CA22556 (N.A.N.), the Victorian State Government Operational Infrastructure Support grant and the NHMRC Independent Research Institutes Infrastructure Support Scheme (361646). N.A.N. acknowledges fellowship support from the NHMRC, J.M.M. and J.J.B. from the Australian Research Council and L.N.V. from the Leukaemia Foundation of Australia and the Australian Stem Cell Centre. We thank the scientists at the MX and SAXS/WAXS beamlines at the Australian Synchrotron. Crystallization trials were performed at CSIRO collaborative crystallization centre (C3).

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Contributions

N.J.K. carried out crystallographic data collection, structure determination and refinement. J.J.B. and N.P.D.L. performed inhibition and co-precipitation assays. J.M.M. and L.N.V. carried out SAXS data collection and analysis. J.M.M., A.L., I.S.L. and E.L.W. performed protein expression and purification experiments. All authors commented on the manuscript . J.J.B., N.J.K. and N.A.N. designed experiments, analyzed data, supervised the project and wrote the paper.

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Correspondence to Nicos A Nicola or Jeffrey J Babon.

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N.A.N. is a founder and member of the scientific advisory board of MuriGen Pty Ltd.

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Kershaw, N., Murphy, J., Liau, N. et al. SOCS3 binds specific receptor–JAK complexes to control cytokine signaling by direct kinase inhibition. Nat Struct Mol Biol 20, 469–476 (2013). https://doi.org/10.1038/nsmb.2519

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