RESUMO
Ciliopathies are a large group of clinically and genetically heterogeneous disorders caused by defects in primary cilia. Here we identified mutations in TRAF3IP1 (TNF Receptor-Associated Factor Interacting Protein 1) in eight patients from five families with nephronophthisis (NPH) and retinal degeneration, two of the most common manifestations of ciliopathies. TRAF3IP1 encodes IFT54, a subunit of the IFT-B complex required for ciliogenesis. The identified mutations result in mild ciliary defects in patients but also reveal an unexpected role of IFT54 as a negative regulator of microtubule stability via MAP4 (microtubule-associated protein 4). Microtubule defects are associated with altered epithelialization/polarity in renal cells and with pronephric cysts and microphthalmia in zebrafish embryos. Our findings highlight the regulation of cytoplasmic microtubule dynamics as a role of the IFT54 protein beyond the cilium, contributing to the development of NPH-related ciliopathies.
Assuntos
Proteínas de Transporte/genética , Doenças Renais Císticas/genética , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Microtúbulos/metabolismo , Mutação , Degeneração Retiniana/genética , Proteínas de Peixe-Zebra/genética , Animais , Western Blotting , Proteínas de Transporte/metabolismo , Polaridade Celular/genética , Dicroísmo Circular , Embrião não Mamífero , Feminino , Imunofluorescência , Técnicas de Inativação de Genes , Células HEK293 , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Imunoprecipitação , Doenças Renais Císticas/metabolismo , Masculino , Microftalmia/genética , Linhagem , Degeneração Retiniana/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Peixe-Zebra , Proteínas de Peixe-Zebra/metabolismoRESUMO
Nephronophthisis is an autosomal recessive cystic kidney disease that leads to renal failure in childhood or adolescence. Most NPHP gene products form molecular networks. Here we identify ANKS6 as a new NPHP family member that connects NEK8 (NPHP9) to INVS (NPHP2) and NPHP3. We show that ANKS6 localizes to the proximal cilium and confirm its role in renal development through knockdown experiments in zebrafish and Xenopus laevis. We also identify six families with ANKS6 mutations affected by nephronophthisis, including severe cardiovascular abnormalities, liver fibrosis and situs inversus. The oxygen sensor HIF1AN hydroxylates ANKS6 and INVS and alters the composition of the ANKS6-INVS-NPHP3 module. Knockdown of Hif1an in Xenopus results in a phenotype that resembles loss of other NPHP proteins. Network analyses uncovered additional putative NPHP proteins and placed ANKS6 at the center of this NPHP module, explaining the overlapping disease manifestation caused by mutation in ANKS6, NEK8, INVS or NPHP3.
Assuntos
Doenças Renais Císticas/genética , Cinesinas/genética , Proteínas Nucleares/genética , Proteínas Quinases/genética , Fatores de Transcrição/genética , Animais , Cílios/metabolismo , Consanguinidade , Éxons , Técnicas de Silenciamento de Genes , Humanos , Íntrons , Doenças Renais Císticas/metabolismo , Cinesinas/metabolismo , Camundongos , Mutação , Quinases Relacionadas a NIMA , Proteínas Nucleares/metabolismo , Fenótipo , Doenças Renais Policísticas/genética , Ligação Proteica , Mapas de Interação de Proteínas , Proteínas Quinases/metabolismo , Transporte Proteico , Fatores de Transcrição/metabolismo , Xenopus/embriologia , Xenopus/metabolismo , Peixe-Zebra/genética , Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismoRESUMO
BACKGROUND: Congenital nephrotic syndrome (CNS) is defined as nephrotic syndrome that manifests within the first 3 months of life. Mutations in the NPHS1 gene encoding nephrin, are a major cause for CNS. Currently, more than 173 different mutations of NPHS1 have been published as causing CNS, affecting most exons. METHODS: We performed mutation analysis of NPHS1 in a worldwide cohort of 20 families (23 children) with CNS. All 29 exons of the NPHS1 gene were examined using direct sequencing. New mutations were confirmed by demonstrating their absence in 96 healthy control individuals. RESULTS: We detected disease-causing mutations in 9 of 20 families (45%). Seven of the families showed a homozygous mutation, while two were compound heterozygous. In another 2 families, single heterozygous NPHS1 mutations were detected. Out of 10 different mutations discovered, 3 were novel, consisting of 1 splice site mutation and 2 missense mutations. CONCLUSION: Our data demonstrate that the spectrum of NPHS1 mutations is still expanding, involving new exons, in patients from a diverse ethnic background.
