Super-resolution microscopy reveals that disruption of ciliary transition-zone architecture causes Joubert syndrome

Ciliopathies, including nephronophthisis (NPHP), Meckel syndrome (MKS) and Joubert syndrome (JBTS), can be caused by mutations affecting components of the transition zone, a domain near the base of the cilium that controls the protein composition of its membrane. We defined the three-dimensional arrangement of key proteins in the transition zone using two-colour stochastic optical reconstruction microscopy (STORM). NPHP and MKS complex components form nested rings comprised of nine-fold doublets. JBTS-associated mutations in RPGRIP1L or TCTN2 displace certain transition-zone proteins. Diverse ciliary proteins accumulate at the transition zone in wild-type cells, suggesting that the transition zone is a waypoint for proteins entering and exiting the cilium. JBTS-associated mutations in RPGRIP1L disrupt SMO accumulation at the transition zone and the ciliary localization of SMO. We propose that the disruption of transition-zone architecture in JBTS leads to a failure of SMO to accumulate at the transition zone and cilium, disrupting developmental signalling in JBTS.

[1]  Christian Cole,et al.  JPred4: a protein secondary structure prediction server , 2015, Nucleic Acids Res..

[2]  Nansheng Chen,et al.  MKS and NPHP modules cooperate to establish basal body/transition zone membrane associations and ciliary gate function during ciliogenesis , 2011, The Journal of cell biology.

[3]  G. Pazour,et al.  NPHP4 controls ciliary trafficking of membrane proteins and large soluble proteins at the transition zone , 2014, Journal of Cell Science.

[4]  D. Storm,et al.  Adult Type 3 Adenylyl Cyclase–Deficient Mice Are Obese , 2009, PloS one.

[5]  Mark Bates,et al.  Multicolor Super-Resolution Imaging with Photo-Switchable Fluorescent Probes , 2007, Science.

[6]  R. Salomon,et al.  Mutational analysis of the RPGRIP1L gene in patients with Joubert syndrome and nephronophthisis. , 2007, Kidney international.

[7]  Manuel Guizar-Sicairos,et al.  Efficient subpixel image registration algorithms. , 2008, Optics letters.

[8]  S. Brody,et al.  Culture and differentiation of mouse tracheal epithelial cells. , 2013, Methods in molecular biology.

[9]  M. Scott,et al.  Patched 1 Regulates Hedgehog Signaling at the Primary Cilium , 2022 .

[10]  G. C. Rogers,et al.  Subdiffraction-resolution fluorescence microscopy reveals a domain of the centrosome critical for pericentriolar material organization , 2012, Nature Cell Biology.

[11]  Mark Bates,et al.  Three-Dimensional Super-Resolution Imaging by Stochastic Optical Reconstruction Microscopy , 2008, Science.

[12]  N. Gilula,et al.  THE CILIARY NECKLACE , 1972, The Journal of cell biology.

[13]  Colin A. Johnson,et al.  The ciliary gene RPGRIP1L is mutated in cerebello-oculo-renal syndrome (Joubert syndrome type B) and Meckel syndrome , 2007, Nature Genetics.

[14]  Ben M. Webb,et al.  Comparative Protein Structure Modeling Using MODELLER , 2007, Current protocols in protein science.

[15]  J. Reiter,et al.  Scoring a backstage pass: Mechanisms of ciliogenesis and ciliary access , 2012, The Journal of cell biology.

[16]  Laurence Pelletier,et al.  Subdiffraction imaging of centrosomes reveals higher-order organizational features of pericentriolar material , 2012, Nature Cell Biology.

[17]  Richard G. W. Anderson,et al.  THE THREE-DIMENSIONAL STRUCTURE OF THE BASAL BODY FROM THE RHESUS MONKEY OVIDUCT , 1972, The Journal of cell biology.

[18]  Arne Elofsson,et al.  The TOPCONS web server for consensus prediction of membrane protein topology and signal peptides , 2015 .

[19]  Colin A. Johnson,et al.  Unraveling the genetics of Joubert and Meckel-Gruber syndromes , 2014, Journal of Pediatric Genetics.

[20]  Kevin W Eliceiri,et al.  NIH Image to ImageJ: 25 years of image analysis , 2012, Nature Methods.

[21]  V. Torres,et al.  B9D1 is revealed as a novel Meckel syndrome (MKS) gene by targeted exon-enriched next-generation sequencing and deletion analysis. , 2011, Human molecular genetics.

[22]  H. Zentgraf,et al.  Mutations in KIF7 link Joubert syndrome with Sonic Hedgehog signaling and microtubule dynamics. , 2011, The Journal of clinical investigation.

[23]  R. Sudbrak,et al.  A gene mutated in nephronophthisis and retinitis pigmentosa encodes a novel protein, nephroretinin, conserved in evolution. , 2002, American journal of human genetics.

[24]  M. Noll,et al.  The Drosophila smoothened Gene Encodes a Seven-Pass Membrane Protein, a Putative Receptor for the Hedgehog Signal , 1996, Cell.

[25]  A. Ashley-Koch,et al.  TMEM231, mutated in orofaciodigital and Meckel syndromes, organizes the ciliary transition zone , 2015, The Journal of cell biology.

[26]  Gabriel Taubin,et al.  Estimation of Planar Curves, Surfaces, and Nonplanar Space Curves Defined by Implicit Equations with Applications to Edge and Range Image Segmentation , 1991, IEEE Trans. Pattern Anal. Mach. Intell..

[27]  J. Liao,et al.  Superresolution Pattern Recognition Reveals the Architectural Map of the Ciliary Transition Zone , 2015, Scientific Reports.

[28]  W E Moerner,et al.  STED microscopy with optimized labeling density reveals 9-fold arrangement of a centriole protein. , 2012, Biophysical journal.

[29]  F. Andermann,et al.  Familial Agenesis of the Cerebellar Vermis: A Syndrome of Episodic Hyperpnea, Abnormal Eye Movements, Ataxia, and Retardation , 1999, Neurology.

[30]  R. Myers,et al.  Human Homolog of patched, a Candidate Gene for the Basal Cell Nevus Syndrome , 1996, Science.

[31]  W. Chung,et al.  Hypomorphism for RPGRIP1L, a ciliary gene vicinal to the FTO locus, causes increased adiposity in mice. , 2014, Cell metabolism.

[32]  J. Reiter,et al.  Conserved Genetic Interactions between Ciliopathy Complexes Cooperatively Support Ciliogenesis and Ciliary Signaling , 2015, PLoS genetics.

[33]  Valerio Gennaro.  [The power of numbers]. , 2005, Epidemiologia e prevenzione.

[34]  C. Pearson,et al.  Basal Body Assembly in Ciliates: The Power of Numbers , 2009, Traffic.

[35]  M. Scott,et al.  Altered neural cell fates and medulloblastoma in mouse patched mutants. , 1997, Science.

[36]  D. Storm,et al.  Inactivation of the mouse adenylyl cyclase 3 gene disrupts male fertility and spermatozoon function. , 2005, Molecular endocrinology.

[37]  J. Reiter,et al.  The base of the cilium: roles for transition fibres and the transition zone in ciliary formation, maintenance and compartmentalization , 2012, EMBO reports.

[38]  S. Dutcher,et al.  The awesome power of dikaryons for studying flagella and basal bodies in Chlamydomonas reinhardtii , 2014, Cytoskeleton.

[39]  Lucien E. Weiss,et al.  Single-molecule imaging of Hedgehog pathway protein Smoothened in primary cilia reveals binding events regulated by Patched1 , 2015, Proceedings of the National Academy of Sciences.

[40]  Michael Dean,et al.  Mutations of the Human Homolog of Drosophila patched in the Nevoid Basal Cell Carcinoma Syndrome , 1996, Cell.

[41]  J. B. Rattner,et al.  Adenylate cyclase regulates elongation of mammalian primary cilia. , 2009, Experimental cell research.

[42]  Carsten Bergmann,et al.  Disruption of a ciliary B9 protein complex causes Meckel syndrome. , 2011, American journal of human genetics.

[43]  R. Dildrop,et al.  Ftm is a novel basal body protein of cilia involved in Shh signalling , 2007, Development.

[44]  J. Rosenbaum,et al.  CEP290 tethers flagellar transition zone microtubules to the membrane and regulates flagellar protein content , 2010, The Journal of cell biology.

[45]  Matthew J. Brauer,et al.  Mapping the NPHP-JBTS-MKS Protein Network Reveals Ciliopathy Disease Genes and Pathways , 2011, Cell.

[46]  J. García-Verdugo,et al.  A Transition Zone Complex Regulates Mammalian Ciliogenesis and Ciliary Membrane Composition , 2011, Nature Genetics.

[47]  I. Glass,et al.  Mutations in the gene encoding the basal body protein RPGRIP1L, a nephrocystin-4 interactor, cause Joubert syndrome , 2007, Nature Genetics.

[48]  S. Doxsey,et al.  Human basal body basics , 2016, Cilia.

[49]  Ben M. Webb,et al.  Comparative Protein Structure Modeling Using Modeller , 2006, Current protocols in bioinformatics.

[50]  K. Anderson,et al.  The primary cilium: a signalling centre during vertebrate development , 2010, Nature Reviews Genetics.

[51]  C. James,et al.  Sporadic medulloblastomas contain PTCH mutations. , 1997, Cancer research.

[52]  J. Rosenbaum,et al.  Localization of intraflagellar transport protein IFT52 identifies basal body transitional fibers as the docking site for IFT particles , 2001, Current Biology.

[53]  C. Walsh,et al.  Abnormal cerebellar development and axonal decussation due to mutations in AHI1 in Joubert syndrome , 2004, Nature Genetics.

[54]  Alexey Drozdetskiy,et al.  JPred4: a protein secondary structure prediction server , 2015, Nucleic Acids Res..

[55]  X. Zhuang,et al.  Superresolution Imaging of Chemical Synapses in the Brain , 2010, Neuron.

[56]  Jussi Taipale,et al.  Small molecule modulation of Smoothened activity , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[57]  G. Bishop,et al.  Type III adenylyl cyclase localizes to primary cilia throughout the adult mouse brain , 2007, The Journal of comparative neurology.

[58]  Ben M. Webb,et al.  ModBase, a database of annotated comparative protein structure models and associated resources , 2013, Nucleic Acids Res..

[59]  D. Labuda,et al.  Mutations in TMEM231 cause Joubert syndrome in French Canadians , 2012, Journal of Medical Genetics.

[60]  V. Jensen,et al.  Formation of the transition zone by Mks5/Rpgrip1L establishes a ciliary zone of exclusion (CIZE) that compartmentalises ciliary signalling proteins and controls PIP2 ciliary abundance , 2015, The EMBO journal.

[61]  J. Reiter,et al.  A primer on the mouse basal body , 2016, Cilia.

[62]  N. Daigle,et al.  Nuclear Pore Scaffold Structure Analyzed by Super-Resolution Microscopy and Particle Averaging , 2013, Science.

[63]  P. Thelwall,et al.  Murine Joubert syndrome reveals Hedgehog signaling defects as a potential therapeutic target for nephronophthisis , 2014, Proceedings of the National Academy of Sciences.

[64]  H. Kayserili,et al.  Mutations in the AHI1 gene, encoding jouberin, cause Joubert syndrome with cortical polymicrogyria. , 2004, American journal of human genetics.

[65]  M. Liebau,et al.  Ciliopathies - from rare inherited cystic kidney diseases to basic cellular function , 2015, Molecular and Cellular Pediatrics.

[66]  Michael W. Davidson,et al.  Nanoscale architecture of integrin-based cell adhesions , 2010, Nature.

[67]  Christian Eggeling,et al.  Multicolor far-field fluorescence nanoscopy through isolated detection of distinct molecular species. , 2008, Nano letters.

[68]  Colin A. Johnson,et al.  Mutations in the cilia gene ARL13B lead to the classical form of Joubert syndrome. , 2008, American journal of human genetics.

[69]  I. Glass,et al.  The NPHP1 gene deletion associated with juvenile nephronophthisis is present in a subset of individuals with Joubert syndrome. , 2004, American journal of human genetics.

[70]  Scott T. Wong,et al.  Disruption of the Type III Adenylyl Cyclase Gene Leads to Peripheral and Behavioral Anosmia in Transgenic Mice , 2000, Neuron.

[71]  Ben Chih,et al.  A ciliopathy complex at the transition zone protects the cilia as a privileged membrane domain , 2011, Nature Cell Biology.

[72]  Didier Y. R. Stainier,et al.  Vertebrate Smoothened functions at the primary cilium , 2005, Nature.