Disease‐causing mutations in the XIAP BIR 2 domain impair NOD 2‐dependent immune signalling

Rune Busk Damgaard,Berthe Katrine Fiil,Monica Yabal,Simon Bekker‐Jensen,Philipp J Jost,Mads Gyrd‐Hansen,Niels Mailand,Stephan Ehl,Udo Zur Stadt,Carsten Speckmann

doi:10.1002/emmm.201303090

Abstract

X-linked Inhibitor of Apoptosis (XIAP) is an essential ubiquitin ligase for pro-inflammatory signalling downstream of the nucleotide-binding oligomerization domain containing (NOD)-1 and -2 pattern recognition receptors. Mutations in XIAP cause X-linked lymphoproliferative syndrome type-2 (XLP2), an immunodeficiency associated with a potentially fatal deregulation of the immune system, whose aetiology is not well understood. Here, we identify the XIAP baculovirus IAP repeat (BIR)2 domain as a hotspot for missense mutations in XLP2. We demonstrate that XLP2-BIR2 mutations severely impair NOD1/2-dependent immune signalling in primary cells from XLP2 patients and in reconstituted XIAP-deficient cell lines. XLP2-BIR2 mutations abolish the XIAP-RIPK2 interaction resulting in impaired ubiquitylation of RIPK2 and recruitment of linear ubiquitin chain assembly complex (LUBAC) to the NOD2-complex. We show that the RIPK2 binding site in XIAP overlaps with the BIR2 IBM-binding pocket and find that a bivalent Smac mimetic compound (SMC) potently antagonises XIAP function downstream of NOD2 to limit signalling. These findings suggest that impaired immune signalling in response to NOD1/2 stimulation is a general defect in XLP2 and demonstrate that the XIAP BIR2-RIPK2 interaction may be targeted pharmacologically to modulate inflammatory signalling.The X-linked lymphoproliferative syndrome type-2 is an immunodeficiency disease caused by mutations in the XIAP gene. BIR2 domain mutations in patients impair RIPK2 binding and NOD2-dependent innate immune signaling, explaining some of the pathology.

Highlights

Disease‐causing mutations in X-linked Inhibitor of Apoptosis (XIAP)/BIRC4 were first described in 2006 in families with patients suffering from X‐linked lymphoproliferative syndrome (XLP) with no mutations in the SH2D1A gene encoding SAP (Rigaud et al, 2006)
We report here three novel missense mutations identified in X-linked lymphoproliferative syndrome type-2 (XLP2) patients, c.497G > T (p.R166I), c.620T > C (p.L207P) and c.592G > A (p.V198M), all locating to the BIR2 domain of XIAP [Fig 1A; during preparation of this manuscript the R166I mutations has been reported by others (Marsh et al, 2013)]
We find that RIPK2 binding and NOD2 signalling is antagonized by mutations in the XIAP BIR2 IAP binding motif (IBM)‐binding pocket, expression of Second mitochondria‐derived activator of caspases (Smac) or incubation with Smac mimetic compound (SMC), Compound A (CpA) in particular

Summary

Introduction

Disease‐causing mutations in XIAP/BIRC4 were first described in 2006 in families with patients suffering from X‐linked lymphoproliferative syndrome (XLP) with no mutations in the SH2D1A gene encoding SAP (Rigaud et al, 2006). Classical XLP due to SAP deficiency (XLP1) is characterized by susceptibility to (1) Department of Disease Biology, Novo Nordisk Foundation Center for Protein Research, University of Copenhagen, Copenhagen, Denmark (2) CCI – Centre of Chronic Immunodeficiency, University Hospital Freiburg, Freiburg, Germany (3) III. XLP2 caused by mutation in XIAP shares the susceptibility to EBV with a high risk of HLH, but no patient with lymphoma has so far been reported (Filipovich et al, 2010; Pachlopnik Schmid et al, 2011; Yang et al, 2012). Severe chronic colitis, hepatitis or persistent splenomegaly are increasingly reported as initial and even as the only clinical manifestations of patients with XIAP mutations [(Worthey et al, 2011), Carsten Speckmann et al, in preparation]. The molecular basis of these inflammatory manifestations remains poorly characterized

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