Shichen Sun1, Fang Li1, Yang Liu1, Hong Qu2, Sing-Wai Wong3, Li Zeng1, Miao Yu1, Hailan Feng1, Haochen Liu4, Dong Han5. 1. Department of Prosthodontics, Peking University School and Hospital of Stomatology & National Clinical Research Center for Oral Diseases & National Engineering Laboratory for Digital and Material Technology of Stomatology & Beijing Key Laboratory of Digital Stomatology, 22 Zhongguancun South Avenue, Haidian District, Beijing 100081, PR China. 2. Center for Bioinformatics, State Key Laboratory of Protein and Plant Gene Research, College of Life Sciences, Peking University, Beijing 100871, PR China. 3. Oral and Craniofacial Biomedicine Curriculum, School of Dentistry, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA. 4. Department of Prosthodontics, Peking University School and Hospital of Stomatology & National Clinical Research Center for Oral Diseases & National Engineering Laboratory for Digital and Material Technology of Stomatology & Beijing Key Laboratory of Digital Stomatology, 22 Zhongguancun South Avenue, Haidian District, Beijing 100081, PR China. Electronic address: talentlhc@163.com. 5. Department of Prosthodontics, Peking University School and Hospital of Stomatology & National Clinical Research Center for Oral Diseases & National Engineering Laboratory for Digital and Material Technology of Stomatology & Beijing Key Laboratory of Digital Stomatology, 22 Zhongguancun South Avenue, Haidian District, Beijing 100081, PR China. Electronic address: donghan@bjmu.edu.cn.
Abstract
OBJECTIVE: To explore the gene mutation in an incontinentia pigmenti (IP) patient with syndromic tooth agenesis. METHODS: Long-range polymerase chain reaction (PCR) and Sanger sequencing were used to detect inhibitor of nuclear factor kappa-B kinase subunit gamma (IKBKG) mutation in the IP patient. We used the nuclear factor kappa B (NF-κB) reporter gene to assess activation of NF-κB, after transfecting an empty vector, wild-type, or mutant NF-κB essential modulator (NEMO) plasmid into IKBKG-deficient HEK293T cells, respectively. Furthermore, we performed immunoprecipitation and immunoblotting to describe the polyubiquitination of NEMO. Lastly, we detected the interactions between mutant NEMO and I kappa B kinase alpha (IKKα), I kappa B kinase beta (IKKβ), TNF receptor associated factor 6 (TRAF6), HOIL-1-interacting protein (HOIP), hemo-oxidized iron regulatory protein 2 ligase 1 (HOIL-1), and SHANK-associated RH domain interactor (SHARPIN). RESULTS: A de novo nonsense mutation in IKBKG (c.924C > G; p.Tyr308*) was observed. The Tyr308* mutation inhibited activation of the NF-κB pathway by reducing K63-linked polyubiquitination and linear polyubiquitination. The mutant NEMO was not able to interact with TRAF6, HOIL-1, or SHARPIN. CONCLUSIONS: We identified a novel nonsense IKBKG mutation (c.924C > G; p.Tyr308*) in an IP patient with syndromic tooth agenesis. This research enriches the mutation spectrum of the IKBKG gene.
OBJECTIVE: To explore the gene mutation in an incontinentia pigmenti (IP) patient with syndromic tooth agenesis. METHODS: Long-range polymerase chain reaction (PCR) and Sanger sequencing were used to detect inhibitor of nuclear factor kappa-B kinase subunit gamma (IKBKG) mutation in the IP patient. We used the nuclear factor kappa B (NF-κB) reporter gene to assess activation of NF-κB, after transfecting an empty vector, wild-type, or mutant NF-κB essential modulator (NEMO) plasmid into IKBKG-deficient HEK293T cells, respectively. Furthermore, we performed immunoprecipitation and immunoblotting to describe the polyubiquitination of NEMO. Lastly, we detected the interactions between mutant NEMO and I kappa B kinase alpha (IKKα), I kappa B kinase beta (IKKβ), TNF receptor associated factor 6 (TRAF6), HOIL-1-interacting protein (HOIP), hemo-oxidized iron regulatory protein 2 ligase 1 (HOIL-1), and SHANK-associated RH domain interactor (SHARPIN). RESULTS: A de novo nonsense mutation in IKBKG (c.924C > G; p.Tyr308*) was observed. The Tyr308* mutation inhibited activation of the NF-κB pathway by reducing K63-linked polyubiquitination and linear polyubiquitination. The mutant NEMO was not able to interact with TRAF6, HOIL-1, or SHARPIN. CONCLUSIONS: We identified a novel nonsense IKBKG mutation (c.924C > G; p.Tyr308*) in an IP patient with syndromic tooth agenesis. This research enriches the mutation spectrum of the IKBKG gene.