SHARPIN forms a linear ubiquitin ligase complex regulating NF-κB activity and apoptosis

Abstract

SHARPIN is a ubiquitin-binding and ubiquitin-like-domain-containing protein which, when mutated in mice, results in immune system disorders and multi-organ inflammation1,2. Here we report that SHARPIN functions as a novel component of the linear ubiquitin chain assembly complex (LUBAC) and that the absence of SHARPIN causes dysregulation of NF-κB and apoptotic signalling pathways, explaining the severe phenotypes displayed by chronic proliferative dermatitis (cpdm) in SHARPIN-deficient mice. Upon binding to the LUBAC subunit HOIP (also known as RNF31), SHARPIN stimulates the formation of linear ubiquitin chains in vitro and in vivo. Coexpression of SHARPIN and HOIP promotes linear ubiquitination of NEMO (also known as IKBKG), an adaptor of the IκB kinases (IKKs) and subsequent activation of NF-κB signalling, whereas SHARPIN deficiency in mice causes an impaired activation of the IKK complex and NF-κB in B cells, macrophages and mouse embryonic fibroblasts (MEFs). This effect is further enhanced upon concurrent downregulation of HOIL-1L (also known as RBCK1), another HOIP-binding component of LUBAC. In addition, SHARPIN deficiency leads to rapid cell death upon tumour-necrosis factor α (TNF-α) stimulation via FADD- and caspase-8-dependent pathways. SHARPIN thus activates NF-κB and inhibits apoptosis via distinct pathways in vivo.

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Figure 1: SHARPIN is a novel component of the LUBAC complex.
Figure 2: SHARPIN and HOIP form a novel LUBAC complex with the ability to induce linear ubiquitination and NF-κB activation.
Figure 3: SHARPIN and HOIL-1L are essential for full activation of IKK and NF-κB.
Figure 4: Loss of SHARPIN sensitizes cells to FADD- and caspase-8-mediated apoptosis

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Acknowledgements

We thank E. Kim, K. Rajalingham and H.-J. Kreienkampfor reagents used in this study, I. Matic for initial MS analysis of HOIP/HOIL-1L samples, S. Wahl for sample preparation, and V. Dötsch and members of the Dikic lab for discussions and comments. This work was supported by grants from the Deutsche Forschungsgemeinschaft (DI 931/3-1), the Cluster of Excellence “Macromolecular Complexes” of the Goethe University Frankfurt (EXC115) to I.D., Landesstiftung Baden-Württemberg to B.M., the Medical Research Council UK to K.R. and B.S., JSPS Postdoctoral Fellowships for Research Abroad to F.I., EMBO long-term fellowship to S.S.S. and The National Institutes of Health (AR049288 to J.P.S.). V.N. was supported by the Unity Through Knowledge Fund, 3B Grant. C.G. acknowledges support from The International Human Frontier Science Program Organization.

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F.I., Y.L.D., S.S.S., B.S., C.G., S.J.L.v.W., B.M., V.N., M.F.-W. and P.G. performed the experiments. F.T., A.A. and T.N. contributed with reagents used throughout the study. F.I., Y.L.D., S.S.S., C.G., M.P., J.T., K.I., J.P.S., L.F., B.M. and K.R. contributed to the project by co-ordination of experimental work and writing the manuscript. I.D. provided ideas, co-ordinated the entire project and wrote the manuscript.

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Correspondence to Ivan Dikic.

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Ikeda, F., Deribe, Y., Skånland, S. et al. SHARPIN forms a linear ubiquitin ligase complex regulating NF-κB activity and apoptosis. Nature 471, 637–641 (2011). https://doi.org/10.1038/nature09814

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