Human hypomorphic NEMO mutations cause diverse clinical immunologic phenotypes, but understanding their scope and mechanistic links immune function and genotype is incomplete.

We created and analyzed a database of hypomorphic NEMO mutations to determine the spectrum of phenotypes and their associated genotypes and sought to establish a standardized NEMO reconstitution system to obtain mechanistic insights.

Phenotypes of 72 individuals with NEMO mutations were compiled. NEMO L153R and C417R were investigated further in a reconstitution system. TNF-α or Toll-like receptor 5 signals were evaluated for NF-κB activation, programmed cell death, and A20 gene expression.

32 different mutations were identified; 53% affect the zinc finger domain. 81% were associated with Ectodermal dysplasia, 76% with serious pyogenic infection, 39% with mycobacterial infection, 19% with serious viral infection, 21% with inflammatory diseases. 36% died at a mean age of 6.4 years. CD40, IL-1, TNF-α, TLR, and TCR signals were impaired in 15/16 (94%), 6/7 (86%), 9/11 (77%), 9/14 (64%), and 7/18 (39%), respectively. Hypomorphism-reconstituted NEMO-deficient cells demonstrated partial restoration of NEMO functions. Although both L153R and C417R impaired TLR and TNF-α induced NF-κB activation, L153R also increased TNF-α-induced programmed cell death with decreased A20 expression.

Distinct NEMO hypomorphs define specific disease and genetic characteristics. A reconstitution system can identify attributes of hypomorphisms independent of an individual’s genetic background. Apoptosis susceptibility in L153R reconstituted cells defines a specific phenotype of this mutation that likely contributes to the excessive inflammation with which it is clinically associated.