P085 Skewed binding of linear ubiquitin chains underlies inflammatory pathology in NEMO syndrome

Abstract

Regulated NF- κ B activation during development and the immune response is controlled by NEMO, the NF- κ B essential modulator. Mutations in the IKBG gene encoding NEMO that result in loss of function in NEMO result in the syndrome of Ectodermal Dysplasia with Anhidrosis and Immunodeficiency (EDA-ID), also referred to as NEMO syndrome. By detailed functional analysis of primary patient cells and a NEMO deficient T cell line reconstituted with mutant forms of NEMO we describe a distinct group of NEMO mutations with C-terminal deletion (ΔCT NEMO) which result in EDA-ID combined severe skin and intestinal inflammatory disease. T cells reconstituted with ΔCT NEMO mutations exhibited increased constitutive NF- κ Bactivity, and increased activation following TNF or Flagellin stimulation due to upregulated intrinsic IKK complex activity, but were unresponsive to TCR stimulation. ΔCT NEMO mutations reduced NEMO/IKK complex interactions with K63- but not linear (also referred to M1) polyubiquitin chains. Skewed M1/K63 polyubiquitin binding by ΔCT NEMO led to increased association with RIP1 and stabilization of RIP1 within the NEMO signaling complex. These gain-of-function phenotypes were blocked by mutations targeting the NEMO residues in the UBAN domain essential for binding to linear but not K63-linked polyubiquitin chains. Our findings reveal altered recognition of polyubiquitin modifications by NEMO as a receptor-specific control switch for IKK activity, and identifies the NEMO C-terminus as a negative regulator of induced NF- κ B activation by innate but not adaptive immune receptors.