title: Novel NEK8 Mutations Cause Severe Syndromic Renal Cystic Dysplasia through YAP Dysregulation.
abstract: Ciliopathies are a group of genetic multi-systemic disorders related to dysfunction of the primary cilium, a sensory organelle present at the cell surface that regulates key signaling pathways during development and tissue homeostasis. In order to identify novel genes whose mutations would cause severe developmental ciliopathies, >500 patients/fetuses were analyzed by a targeted high throughput sequencing approach allowing exome sequencing of >1200 ciliary genes. NEK8/NPHP9 mutations were identified in five cases with severe overlapping phenotypes including renal cystic dysplasia/hypodysplasia, situs inversus, cardiopathy with hypertrophic septum and bile duct paucity. These cases highlight a genotype-phenotype correlation, with missense and nonsense mutations associated with hypodysplasia and enlarged cystic organs, respectively. Functional analyses of NEK8 mutations in patient fibroblasts and mIMCD3 cells showed that these mutations differentially affect ciliogenesis, proliferation/apoptosis/DNA damage response, as well as epithelial morphogenesis. Notably, missense mutations exacerbated some of the defects due to NEK8 loss of function, highlighting their likely gain-of-function effect. We also showed that NEK8 missense and loss-of-function mutations differentially affect the regulation of the main Hippo signaling effector, YAP, as well as the expression of its target genes in patient fibroblasts and renal cells. YAP imbalance was also observed in enlarged spheroids of Nek8-invalidated renal epithelial cells grown in 3D culture, as well as in cystic kidneys of Jck mice. Moreover, co-injection of nek8 MO with WT or mutated NEK8-GFP RNA in zebrafish embryos led to shortened dorsally curved body axis, similar to embryos injected with human YAP RNA. Finally, treatment with Verteporfin, an inhibitor of YAP transcriptional activity, partially rescued the 3D spheroid defects of Nek8-invalidated cells and the abnormalities of NEK8-overexpressing zebrafish embryos. Altogether, our study demonstrates that NEK8 human mutations cause major organ developmental defects due to altered ciliogenesis and cell differentiation/proliferation through deregulation of the Hippo pathway.
authors.affiliation: INSERM UMR1163, Laboratory of Inherited Kidney Diseases, Necker-Enfants Malades Hospital, Paris, France.|Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.
authors.forename: Valentina
authors.initials: V
authors.lastname: Grampa
authors.affiliation: INSERM UMR1163, Laboratory of Inherited Kidney Diseases, Necker-Enfants Malades Hospital, Paris, France.|Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.
authors.forename: Marion
authors.initials: M
authors.lastname: Delous
authors.affiliation: INSERM U1151, CNRS UMR8253, Paris Descartes-Sorbonne Paris Cité University, Necker-Enfants Malades Institute, Mechanisms and Therapeutic Strategies of Chronic Kidney Diseases, Necker Hospital, Paris, France.
authors.forename: Mohamad
authors.initials: M
authors.lastname: Zaidan
authors.affiliation: INSERM UMR1163, Laboratory of Inherited Kidney Diseases, Necker-Enfants Malades Hospital, Paris, France.|Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.
authors.forename: Gweltas
authors.initials: G
authors.lastname: Odye
authors.affiliation: Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.|INSERM UMR1163, Laboratory of Embryology and Genetics of Congenital Malformations, Paris, France.
authors.forename: Sophie
authors.initials: S
authors.lastname: Thomas
authors.affiliation: Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.|INSERM UMR1163, Laboratory of Embryology and Genetics of Congenital Malformations, Paris, France.|Department of Genetics, AP-HP, Necker Hospital, Paris, France.
authors.forename: Nadia
authors.initials: N
authors.lastname: Elkhartoufi
authors.affiliation: INSERM UMR1163, Laboratory of Inherited Kidney Diseases, Necker-Enfants Malades Hospital, Paris, France.|Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.
authors.forename: Emilie
authors.initials: E
authors.lastname: Filhol
authors.affiliation: INSERM UMR1163, Laboratory of Inherited Kidney Diseases, Necker-Enfants Malades Hospital, Paris, France.|Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.|Department of Pediatric Nephrology, AP-HP, Robert Debré Hospital, Paris, France.
authors.forename: Olivier
authors.initials: O
authors.lastname: Niel
authors.affiliation: INSERM UMR1163, Laboratory of Inherited Kidney Diseases, Necker-Enfants Malades Hospital, Paris, France.|Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.
authors.forename: Flora
authors.initials: F
authors.lastname: Silbermann
authors.affiliation: Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.|INSERM UMR1163, Laboratory of Intestinal Immunity, Paris, France.
authors.forename: Corinne
authors.initials: C
authors.lastname: Lebreton
authors.affiliation: Department of Pathology, Hospices Civils de Lyon, CHU de Lyon, Lyon, France.
authors.forename: Sophie
authors.initials: S
authors.lastname: Collardeau-Frachon
authors.affiliation: Cellular Biotechnology Department and Biobank, Hospices Civils de Lyon, CHU de Lyon, Lyon, France.
authors.forename: Isabelle
authors.initials: I
authors.lastname: Rouvet
authors.affiliation: CHU de La Réunion Saint-Denis/Saint-Pierre, La Réunion, France.
authors.forename: Jean-Luc
authors.initials: JL
authors.lastname: Alessandri
authors.affiliation: Anatomopathological Department, CHRU Lille, University Hospital, Lille, France.
authors.forename: Louise
authors.initials: L
authors.lastname: Devisme
authors.affiliation: Department of Clinical Genetics, CHRU Lille, Lille, France.
authors.forename: Anne
authors.initials: A
authors.lastname: Dieux-Coeslier
authors.affiliation: Department of Genetics, Femme Mère-Enfant Hospital, University of Lyon 1, Bron, France.
authors.forename: Marie-Pierre
authors.initials: MP
authors.lastname: Cordier
authors.affiliation: Department of Genetics, CHU Robert-Debré, Paris, France.
authors.forename: Yline
authors.initials: Y
authors.lastname: Capri
authors.affiliation: Fetal Pathology department, CHU Robert-Debré, Paris, France.
authors.forename: Suonavy
authors.initials: S
authors.lastname: Khung-Savatovsky
authors.affiliation: Multidisciplinary Department of Prenatal Diagnosis, La Timone Children's Hospital, Marseille, France.
authors.forename: Sabine
authors.initials: S
authors.lastname: Sigaudy
authors.affiliation: Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.|Department of Pediatric Nephrology, AP-HP, Necker Hospital, Paris, France.
authors.forename: Rémi
authors.initials: R
authors.lastname: Salomon
authors.affiliation: INSERM UMR1163, Laboratory of Inherited Kidney Diseases, Necker-Enfants Malades Hospital, Paris, France.|Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.|Department of Genetics, AP-HP, Necker Hospital, Paris, France.
authors.forename: Corinne
authors.initials: C
authors.lastname: Antignac
authors.affiliation: INSERM UMR1163, Laboratory of Inherited Kidney Diseases, Necker-Enfants Malades Hospital, Paris, France.|Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.
authors.forename: Marie-Claire
authors.initials: MC
authors.lastname: Gubler
authors.affiliation: INSERM UMR1163, Laboratory of Inherited Kidney Diseases, Necker-Enfants Malades Hospital, Paris, France.|Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.
authors.forename: Alexandre
authors.initials: A
authors.lastname: Benmerah
authors.affiliation: INSERM U1151, CNRS UMR8253, Paris Descartes-Sorbonne Paris Cité University, Necker-Enfants Malades Institute, Mechanisms and Therapeutic Strategies of Chronic Kidney Diseases, Necker Hospital, Paris, France.
authors.forename: Fabiola
authors.initials: F
authors.lastname: Terzi
authors.affiliation: Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.|INSERM UMR1163, Laboratory of Embryology and Genetics of Congenital Malformations, Paris, France.|Department of Histology-Embryology and Cytogenetics, AP-HP, Necker Hospital, Paris, France.
authors.forename: Tania
authors.initials: T
authors.lastname: Attié-Bitach
authors.affiliation: INSERM UMR1163, Laboratory of Inherited Kidney Diseases, Necker-Enfants Malades Hospital, Paris, France.|Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.
authors.forename: Cécile
authors.initials: C
authors.lastname: Jeanpierre
authors.affiliation: INSERM UMR1163, Laboratory of Inherited Kidney Diseases, Necker-Enfants Malades Hospital, Paris, France.|Paris Descartes-Sorbonne Paris Cité University, Imagine Institute, Paris, France.
authors.forename: Sophie
authors.initials: S
authors.lastname: Saunier
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000235:genetics*; D000818:Animals; D002454:Cell Differentiation / Q000235:genetics; D002923:Cilia / Q000235:genetics* / Q000473:pathology; D005260:Female; D056726:Genetic Association Studies; D006801:Humans; D007668:Kidney / Q000378:metabolism / Q000473:pathology; D051379:Mice; D009024:Morphogenesis / Q000235:genetics; D009154:Mutation; D000072379:NIMA-Related Kinases; D010750:Phosphoproteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000235:genetics*; D007690:Polycystic Kidney Diseases / Q000235:genetics* / Q000473:pathology; D011166:Porphyrins / Q000008:administration & dosage; D011494:Protein Kinases / Q000235:genetics*; D015398:Signal Transduction; D014157:Transcription Factors; D000077362:Verteporfin; D000091102:YAP-Signaling Proteins; D015027:Zebrafish
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D010750:Phosphoproteins; D011166:Porphyrins; D014157:Transcription Factors; D000091102:YAP-Signaling Proteins; C088374:YAP1 protein, human; D000077362:Verteporfin; D011494:Protein Kinases; C458529:NEK8 protein, human; D000072379:NIMA-Related Kinases