2. Academic Validation
  3. Functional assessment of the mutational effects of human IRAK4 and MyD88 genes

Functional assessment of the mutational effects of human IRAK4 and MyD88 genes

  • Mol Immunol. 2014 Mar;58(1):66-76. doi: 10.1016/j.molimm.2013.11.008.
Takahiro Yamamoto 1,  Naotaka Tsutsumi 2,  Hidehito Tochio 3,  Hidenori Ohnishi 4,  Kazuo Kubota 1,  Zenichiro Kato 1,  Masahiro Shirakawa 5,  Naomi Kondo 6
Affiliations

Affiliations

  • 1 Department of Pediatrics, Graduate School of Medicine, Gifu University, 1-1 Yanagido, Gifu 501-1194, Japan.
  • 2 Department of Molecular Engineering, Graduate School of Engineering, Kyoto University, Katsura, Nishikyo-ku, Kyoto 615-8510, Japan.
  • 3 Department of Molecular Engineering, Graduate School of Engineering, Kyoto University, Katsura, Nishikyo-ku, Kyoto 615-8510, Japan. Electronic address: [email protected].
  • 4 Department of Pediatrics, Graduate School of Medicine, Gifu University, 1-1 Yanagido, Gifu 501-1194, Japan. Electronic address: [email protected].
  • 5 Department of Molecular Engineering, Graduate School of Engineering, Kyoto University, Katsura, Nishikyo-ku, Kyoto 615-8510, Japan; Core Research for Evolutional Science and Technology, Japan Science and Technology Corporation, 4-1-8 Hon-cho, Kawaguchi, Saitama 332-0012, Japan.
  • 6 Department of Pediatrics, Graduate School of Medicine, Gifu University, 1-1 Yanagido, Gifu 501-1194, Japan; Heisei College of Health Sciences, 180 Kurono, Gifu 501-1131, Japan.
PMID: 24316379 DOI: 10.1016/j.molimm.2013.11.008
Abstract

Human interleukin-1 receptor-associated kinase 4 (IRAK4) deficiency and myeloid differentiating factor 88 (MyD88) deficiency syndromes are two primary immune-deficiency disorders with innate immune defects. Although new genetic variations of IRAK4 and MyD88 have recently been deposited in the single nucleotide polymorphism (SNP) database, the clinical significance of these variants has not yet been established. Therefore, it is important to establish methods for assessing the association of each gene variation with human diseases. Because cell-based assays, western blotting and an NF-κB reporter gene assay, showed no difference in protein expression and NF-κB activity between R12C and wild-type IRAK4, we examined protein-protein interactions of purified recombinant IRAK4 and MyD88 proteins by analytical gel filtration and NMR titration. We found that the variant of IRAK4, R12C, as well as R20W, located in the death domain of IRAK4 and regarded as a SNP, caused a loss of interaction with MyD88. Our studies suggest that not only the loss of protein expression but also the defect of Myddosome formation could cause IRAK4 and MyD88 deficiency syndromes. Moreover a combination of in vitro functional assays is effective for confirming the pathogenicity of mutants found in IRAK4 and MyD88-deficiency patients.

Keywords

DD; Death domain; ELISA; HEK; ID; IRAK; IRAK4; IRAK4-DD; IRAK4-DD+ID; Immune-deficiency; Interleukin-1 receptor-associated kinase; Mal; Mal-TIR; MyD88; MyD88 adaptor-like; MyD88-DD; MyD88-DD+ID; MyD88-TIR; Myddosome; NMR; SNP; TIR domain; TIR domain of Mal; TIR domain of MyD88; TNF receptor associated factor.; TRAF; Toll/interleukin-1 receptor homology domain; WT; death domain; death domain and internal domain of IRAK4; death domain and internal domain of MyD88; death domain of IRAK4; death domain of MyD88; enzyme-linked immunosorbent assay; human embryonic kidney; internal domain; myeloid differentiating factor 88; nuclear magnetic resonance; single nucleotide polymorphism; wild type.

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