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DeLuca, AP(DeLuca, Adam P.); Weed, MC(Weed, Matthew C.); Haas, CM(Haas, Christine M.); Halder, JA(Halder, Jennifer A.); Stone, EM(Stone, Edwin M.) | JAMA Ophthalmology | August 2015 | Vol. 133(8) | pgs. 967-968 | doi:10.1001/jamaophthalmol.2015.1463 The clinical features of a known mendelian disease can occasionally be mimicked by the random co-occurrence of 2 different conditions in the same individual. We report a case in which whole-exome sequencing in a patient previously suspected to have Usher syndrome revealed disease-causing mutations in BBS1 and SLC26A4 . This case illustrates how detailed and accurate clinical data are needed to interpret exome-scale genetic testing results. A man in his late 30s was referred for evaluation of suspected Usher syndrome. As a toddler, he was diagnosed as having sensorineural hearing loss due to a Mondini malformation after computed tomography showed characteristic cochlear abnormalities (audiograms shown in Figure 1 ). A few years later, his parents noted that he began tripping over objects and having trouble seeing at night. Several years later, he was diagnosed as having retinitis pigmentosa and suspected to have Usher syndrome. At that time, his best-corrected visual acuity was 20/60 OD and 20/80 OS. By his early 20s, it had decreased to counting fingers OD and 20/800 OS. Aside from a paternal great-great-aunt with deafness, mutism, and normal vision, there were no other affected family members. To buy article, click here

Michael Danciger, Vickie Heilbron, Yong-Qing Gao, Dan-Yun Zhao, Samuel G. Jacobson, and Debora B. Farber | Mol. Vis. | Volume 2 (10) | 1996 |  molvis.org/molvis/v2/a10/ to Based on average estimates of the prevalence of non-syndromic  retinitis pigmentosa  (RP) at 1/4,000, there are approximately 1.5 million people in the world with this inherited, progressive, degenerative disease of the retinal photoreceptor cells which often leads to blindness. About 50% of these cases are inherited in an autosomal recessive manner (AR). With the approach of screening the exons of candidate genes in large numbers of unrelated ARRP probands, mutations associated with disease have been found in several candidate genes expressed in rod photoreceptors at very low frequency:  RHO , encoding rhodopsin, 1/126 patients screened;  PDE6B (click to continue reading) Based on average estimates of the prevalence of non-syndromic retinitis pigmentosa (RP) at 1/4,000, there are approximately 1.5 million people in the world with this inherited, progressive, degenerative disease of the retinal photoreceptor cells which often leads to blindness (1,2,3). About 50% of these cases are inherited in an autosomal recessive manner (AR). With the approach of screening the exons of candidate genes in large numbers of unrelated ARRP probands, mutations associated with disease have been found in several candidate genes expressed in rod photoreceptors at very low frequency: RHO , encoding rhodopsin, 1/126 patients screened (4); PDE6B , encoding the beta-subunit of rod cGMP-phosphodiesterase, 4/88 patients (5); PDE6A , encoding the alpha-subunit of rod cGMP-phosphodiesterase, 2/173 patients (6); and CNCG , encoding the alpha-subunit of the rod cGMP-gated channel, 3/173 patients (7). With the approach of linkage analysis in large families, two ARRP loci have so far been discovered: 1q31-q32.1 (8,9) and 6p, distal to RDS-peripherin (10). Also, linkage analysis of an informative consanguineous family led to the discovery of a second homozygous RHO mutation (11). ARRP tends to appear most often in small nuclear families that by themselves are not informative enough to yield significant linkage data, and screening all the exons of candidate genes like PDE6A and PDE6B (each with 22 exons) in large numbers of unrelated probands is costly and time consuming. Therefore, we have taken the approach of studying candidate genes in small nuclear ARRP families with a double screening protocol. Linkage analyses of markers close to the loci of the candidate genes are performed first, and any families where a gene locus clearly does not segregate with disease are ruled out from further study of that gene. DNAs of the probands from the remaining families (where the gene locus cannot be ruled out from segregating with disease) are then screened for mutations in the exons of the candidate gene by SSCPE (single strand conformation polymorphism electrophoresis) and DGGE (denaturing gradient gel electrophoresis). Any exonic variants found are sequenced directly and analyzed within the corresponding family to see if they appear to segregate with disease. With this approach we studied 24 families with inherited retinal degenerations (14 with typical RP) for mutations in the genes PDE6B, MYL5 , PDE6C, CNCG , RHO, ROM1 and RDS-peripherin . We have reported two typical ARRP families where the affecteds uniquely inherited compound heterozygous mutations in the PDE6B gene (12). With a similar approach, homozygous mutations also have been found in PDE6B in the affecteds of two other ARRP families (13,14). Click here to read the entire article

Giuseppe Bonapace, Abdul Waheed, Gul N Shah, William S Sly |  Proc National Academy Science  | 2004 Aug 17 | Vol. 101, Issue 33 | 12300-5 | doi: 10.1073/pnas.0404764101 Carbonic anhydrase (CA) IV is a glycosylphosphotidylinositol-anchored enzyme highly expressed on the plasma face of microcapillaries and especially strongly expressed in the choriocapillaris of the human eye. In collaboration with scientists at the University of Cape Town (Rondebosch, South Africa), we recently showed that the R14W mutation in the signal sequence of CA IV, which they identified in patients with the retinitis pigmentosa (RP) 17 form of autosomal dominant RP, results in accumulation of unfolded protein in the endoplasmic reticulum (ER), leading to ER stress, the unfolded protein response, and apoptosis in a large fraction of transfected COS-7 cells expressing mutant, but not wild-type, CA IV. Read more, click here.

Relatively Common Cause of Autosomal Recessive Early-Onset Retinal Degeneration in the Israeli and Palestinian Populations Avigail Beryozkin, Lina Zelinger, Dikla Bandah-Rozenfeld, Anat Harel, Tim A. Strom, Saul Merin, Itay Chowers, Eyal Banin, Dror Sharon | Investigative Ophthalmology & Visual Science | March 2013 | Vol.54 | 2068-2075 | Introduction Mutations in Crumbs homolog 1 (CRB1) are known to cause severe retinal dystrophies, ranging from Leber congenital amaurosis (LCA) to retinitis pigmentosa (RP). (1–7) LCA is the most severe nonsyndromic retinal dystrophy, characterized by blindness or severe visual impairment from birth, nonrecordable electroretinogram (ERG), nystagmus, hypermetropia, sluggish or absent pupillary responses, and oculodigital reflexes. (2,4–6,8) In contrast, RP is considered a milder and more heterogeneous disorder, with a later age of onset. It is characterized by night blindness followed by gradual loss of peripheral vision, progressive degeneration of photoreceptors, and eventually leads to visual impairment of variable severity that in rare cases can result in complete blindness. (9–12) Patients with RP have impaired ERG responses with a rod > cone pattern of injury, and over time suffer characteristic funduscopic findings, including bone spicule–like pigmentary (BSP) changes, attenuation of retinal vessels, and waxy pallor of the optic discs. (9–12) Retinal dystrophies resulting from CRB1 mutations can be accompanied by additional specific features, including relative preservation of the para-arteriolar retinal pigment epithelium (PPRPE) and Coats-like vasculopathy. (1–5,8,13,14) RP with PPRPE is a form of RP characterized by preservation of the RPE that is adjacent to the retinal arterioles, while the rest of the RPE layer degenerates. Coats-like exudative vasculopathy is characterized by abnormal retinal vessels with increased permeability, leading to exudative retinal detachment that often is accompanied by massive subretinal lipid deposits. 3–5,7,13,15 CRB1 is a human homologue of the drosophila transmembrane crumbs protein, and is expressed in the brain and in the inner segments of mammalian photoreceptors. (2,3,7,16–19) The Crumbs protein is implicated in mechanisms that control cell-cell adhesion, intracellular communication and apicobasal cell polarity. For epithelial cells and photoreceptors, separation of their apical and basal compartments is critical for proper development and function of the cells and the tissue, including adhesion and signaling between and within cells.(2,7,13,16–19) Jacobson et al. suggested that CRB1 mutations underlie developmental defects in LCA, including thickening of the retina and lack of distinct layering in the fully developed adult retina.(13) Rashbass et al. postulated that CRB1 has a role in localizing phototransduction proteins to the apical membrane of the photoreceptors. 18 Thus, nonfunctional CRB1 may impede phototransduction, and lead to progressive dystrophy of the photoreceptors and the RPE, resulting in LCA or RP. Read the article References 1. Bujakowska K Audo I Mohand-Said S CRB1 mutations in inherited retinal dystrophies. Hum Mutat . 2011; 33: 306–315. 2. Gosens I den Hollander AI Cremers FP Roepman R. Composition and function of the Crumbs protein complex in the mammalian retina. Exp Eye Res . 2008; 86: 713–726. 3. den Hollander AI ten Brink JB de Kok YJ Mutations in a human homologue of Drosophila crumbs cause retinitis pigmentosa (RP12). Nat Genet . 1999; 23: 217–221. 4. den Hollander AI Heckenlively JR van den Born LI Leber congenital amaurosis and retinitis pigmentosa with Coats-like exudative vasculopathy are associated with mutations in the crumbs homologue 1 (CRB1) gene. Am J Hum Genet . 2001; 69: 198–203. 5. den Hollander AI Davis J van der Velde-Visser SD CRB1 mutation spectrum in inherited retinal dystrophies. Hum Mutat . 2004; 24: 355–369. 6. den Hollander AI Roepman R Koenekoop RK Cremers FP. Leber congenital amaurosis: genes, proteins and disease mechanisms. Prog Retin Eye Res . 2008; 27: 391–419. 7. Richard M Roepman R Aartsen WM Towards understanding CRUMBS function in retinal dystrophies. Hum Mol Genet . 2006; 15 (spec No 2): R235–R243. 8. Simonelli F Ziviello C Testa F Clinical and molecular genetics of Leber's congenital amaurosis: a multicenter study of Italian patients. Invest Ophthalmol Vis Sci . 2007; 48: 4284–4290. 9. Berson EL. Retinitis pigmentosa. The Friedenwald Lecture. Invest Ophthalmol Vis Sci . 1993; 34: 1659–1676. 10. Bhatti MT. Retinitis pigmentosa, pigmentary retinopathies, and neurologic diseases. Curr Neurol Neurosci Rep . 2006; 6: 403–413. 11. Hartong DT Berson EL Dryja TP. Retinitis pigmentosa. Lancet . 2006; 368: 1795–1809. 12.Jacobson SG Roman AJ Aleman TS Normal central retinal function and structure preserved in retinitis pigmentosa. Invest Ophthalmol Vis Sci . 2010; 51: 1079–1085. 13. Jacobson SG Cideciyan AV Aleman TS Crumbs homolog 1 (CRB1) mutations result in a thick human retina with abnormal lamination. Hum Mol Genet . 2003; 12: 1073–1078. 14. Siemiatkowska AM Arimadyo K Moruz LM Molecular genetic analysis of retinitis pigmentosa in Indonesia using genome-wide homozygosity mapping. Mol Vis . 2011; 17: 3013–3024. 15. Cahill M O'Keefe M Acheson R Classification of the spectrum of Coats' disease as subtypes of idiopathic retinal telangiectasis with exudation. Acta Ophthalmol Scand . 2001; 79: 596–602. 16.Davis JA Handford PA Redfield C. The N1317H substitution associated with Leber congenital amaurosis results in impaired interdomain packing in human CRB1 epidermal growth factor-like (EGF) domains. J Biol Chem . 2007; 282: 28807–28814. 17. den Hollander AI Johnson K de Kok YJ CRB1 has a cytoplasmic domain that is functionally conserved between human and Drosophila. Hum Mol Genet . 2001; 10: 2767–2773. 18. Rashbass P Skaer H. Cell polarity: nailing Crumbs to the scaffold. Curr Biol . 2000; 10: R234–R236.

Abstract Retina and retinal pigment epithelium (RPE) belong to the metabolically most active tissues in the human body. Efficient removal of acid load from retina and RPE is a critical function mediated by the choriocapillaris. However, the mechanism by which pH homeostasis is maintained is largely unknown. Here, we show that a functional complex of carbonic anhydrase 4 (CA4) and Na+/bicarbonate co-transporter 1 (NBC1) is specifically expressed in the choriocapillaris and that missense mutations in CA4 linked to autosomal dominant rod–cone dystrophy disrupt NBC1-mediated HCO3−transport. Our results identify a novel pathogenic pathway in which a defect in a functional complex involved in maintaining pH balances, but not expressed in retina or RPE, leads to photoreceptor degeneration. The importance of a functional CA4 for survival of photoreceptors implies that CA inhibitors, which are widely used as medications, particularly in the treatment of glaucoma, may have long-term adverse effects on vision. Read more

Richard J. Davis, Chun-Wei Hsu, Yi-Ting Tsai, Katherine J. Wert, Javier Sancho-Pelluz, Chyuan-Sheng Lin and Stephen H. Tsang | Journal of Neuroscience|  14 Aug 2013 | 33 (33) | 13475-13483 | doi.org/10.1523/JNEUROSCI.0419-13.2013 The third-most common cause of autosomal recessive retinitis pigmentosa (RP) is due to defective cGMP phosphodiesterase-6 (PDE6). Previous work using viral gene therapy on PDE6-mutant mouse models demonstrated photoreceptors can be rescued if administered before degeneration. However, whether visual function can be rescued after degeneration onset has not been addressed. This is a clinically important question, as newly diagnosed patients exhibit considerable loss of rods and cones in their peripheral retinas. We have generated and characterized a tamoxifen inducible Cre-loxP rescue allele, PDE6B. click here to read entire article.

Aisha Pandor, Rajkumar Ramesar, Sharon Prince | J Cell Biochem ​ | 2010 Oct 15 | Vol. 111, Issue 3 | 735-41 |  doi: 10.1002/jcb.22759 Abstract Retinitis pigmentosa is a highly heterogeneous form of inherited blindness which affects more than 1.3 million individuals worldwide. The RP17 form of the disease is caused by an arginine to tryptophan (R14W) mutation in the signal sequence of carbonic anhydrase IV (CAIV). While CAIV is expressed in the choriocapillaries of the eye and renal epithelium, the R14W mutation results in an exclusively ocular phenotype in affected individuals. In order to investigate the mechanism of disease in RP17 and the lack of kidney phenotype, we compared the subcellular localization and post-translational processing of wild-type (WT)- and mutant-CAIV in three cell types. We show using immunocytochemistry that unlike WT CAIV which is transported to the plasma membrane of transfected COS-7 and HT-1080 cells, the R14W mutant CAIV is retained in the endoplasmic reticulum. Western blot analyses further reveal that whereas the WT CAIV is processed to its mature form in both these cell lines, significant levels of the R14W mutant protein remain in its immature form. Importantly, flow cytometry experiments demonstrate that compared to WT CAIV protein, expression of specifically the R14W CAIV results in an S and G2/M cell-cycle block, followed by apoptosis. Interestingly, when the above experiments were repeated in the human embryonic kidney cell line, HEK-293, strikingly different results were obtained. These cells were unaffected by the expression of the R14W mutant CAIV and were able to process the mutant and WT protein equally effectively. This study has important implications for our understanding of the RP17 phenotype. To read full text, click here

Muhammad Imran Khan, Ferry Kersten, Maleeha Azam, Rob Collin, Alamdar Hussain, Syed Tahir-A Shah, Jan Keunen, Hannie Kremer, Frans Cremers, Raheel Qamar, Anneke den Hollander |  Ophthalmology​ | 2011 Jul 1 | Vol.18, Issue 7 |1444-8 | doi: 10.1016/j.ophtha.2010.10.047    Objective: To describe the mutations in the CLRN1 gene in patients from 2 consanguineous Pakistani families diagnosed with autosomal recessive retinitis pigmentosa (arRP). Participants: Affected and unaffected individuals of 2 consanguineous Pakistani families and 90 unaffected controls from the same population. Informed consent was obtained from participants and the protocol was approved by a local institutional review board.

Isosomppi, J., Västinsalo, H., Geller, S. F., Heon, E., Flannery, J. G., & Sankila, E. M. | Molecular Vision | 2009 Sep 8 | Vol. 15 | pgs. 1806-18 | ncbi.nlm.nih.gov/pmc/articles/PMC2742642/ Abstract Purpose Mutations of clarin 1 ( CLRN1 ) cause Usher syndrome type 3 (USH3). To determine the effects of USH3 mutations on CLRN1 function, we examined the cellular distribution and stability of both normal and mutant CLRN1 in vitro. We also searched for novel disease-causing mutations in a cohort of 59 unrelated Canadian and Finnish USH patients. Methods Mutation screening was performed by DNA sequencing. For the functional studies, wild-type (WT) and mutant CLRN1 genes were expressed as hemagglutinin (HA) tagged fusion proteins by transient transfection of BHK-21 cells. Subcellular localization of CLRN1-HA was examined by confocal microscopy. The N-glycosylation status of CLRN1 was studied by using the N-glycosidase F (PNGase F) enzyme and western blotting. Cycloheximide treatment was used to assess the stability of CLRN1 protein. Results We found three previously reported pathogenic mutations, p.A123D, p.N48K, and p.Y176X, and a novel sequence variant, p.L54P, from the studied USH patients. The WT HA-tagged CLRN1 was correctly trafficked to the plasma membrane, whereas mutant CLRN1-HA proteins were mislocalized and retained in the endoplasmic reticulum. PNGase F treatment of CLRN1-HA resulted in an electrophoretic mobility shift consistent with sugar residue cleavage in WT and in all CLRN1 mutants except in p.N48K mutated CLRN1, in which the mutation abolishes the glycosylation site. Inhibition of protein expression with cycloheximide indicated that WT CLRN1-HA remained stable. In contrast, the CLRN1 mutants showed reduced stability. Conclusions WT CLRN1 is a glycoprotein localized to the plasma membrane in transfected BHK-21 cells. Mutant CLRN1 proteins are mislocalized. We suggest that part of the pathogenesis of USH3 may be associated with defective intracellular trafficking as well as decreased stability of mutant CLRN1 proteins. Introduction Usher syndrome (USH) describes a group of autosomal recessive diseases with bilateral sensorineural hearing loss and visual impairment phenotypically similar to retinitis pigmentosa (RP) [ 1 - 4 ]. Prevalence of USH in different populations is estimated to range from 3.5 to 6.2 per 100,000, thus making it the most frequent cause of combined deaf-blindness worldwide [ 5 ]. The condition has been classified into three clinical subtypes (USH1, USH2, and USH3), based on the severity and progression of the hearing impairment, presence or absence of vestibular dysfunction, and the age of onset of RP [ 1 ]. This classification remains in clinical use, although recent progress on the molecular genetics and clinical research of USH has revealed broad genetic and clinical heterogeneity [ 3 , 6 ]. Atypical forms of USH have been identified within all three clinical types, and there is considerable overlap of symptoms among the subtypes. A distinguishing feature of USH3 is the wide spectrum of nonlinear progressive hearing impairment, which ranges from a near normal to a severe audiometric phenotype [ 7 ]. USH3 patients may also have either normal or decreased vestibular responses [ 8 ]. The rate of visual loss in USH3 is similar to other USH subtypes [ 9 ], with the most recent analyses suggesting that retinal degeneration in USH3 progresses more rapidly than in USH2A [ 10 , 11 ]. The variable phenotype may cause USH3 to be under-diagnosed and it may be more prevalent than previously indicated [ 6 ]. To date, nine USH gene products have been identified: the molecular motor myosin VIIa (USH1B) [ 12 ]; the cell adhesion proteins cadherin 23 (USH1D) [ 13 ] and protocadherin 15 (USH1F) [ 14 , 15 ]; the scaffold proteins harmonin (USH1C) [ 16 ], SANS (USH1G) [ 17 ], and whirlin (USH2D) [ 18 ]; the G-protein-coupled 7-transmembrane receptor VLGR1b (USH2C) [ 19 ]; two isoforms of the extracellular matrix connected protein usherin (USH2A) [ 20 , 21 ]; and the four-pass transmembrane domain protein clarin 1 (USH3) [ 22 , 23 ]. There is growing evidence suggesting that these proteins form a network, which is critical for the development and maintenance of the sensorineural cells in the inner ear and the retina [ 3 , 4 , 24 - 28 ]. Click here to read original source article References Petit C. Usher syndrome: from genetics to pathogenesis. Annu Rev Genomics Hum Genet. 2001;2:271–97. doi: 10.1146/annurev.genom.2.1.271. Ahmed ZM, Riazuddin S, Wilcox ER. The molecular genetics of Usher syndrome. Clin Genet. 2003;63:431–44. doi: 10.1034/j.1399-0004.2003.00109.x. Reiners J, Nagel-Wolfrum K, Jurgens K, Marker T, Wolfrum U. Molecular basis of human Usher syndrome: deciphering the meshes of the Usher protein network provides insights into the pathomechanisms of the Usher disease. Exp Eye Res. 2006;83:97–119. doi: 10.1016/j.exer.2005.11.010. Kremer H, van Wijk E, Märker T, Wolfrum U, Roepman R. Usher syndrome: molecular links of pathogenesis, proteins and pathways. Hum Mol Genet. 2006;15:R262–70. doi: 10.1093/hmg/ddl205. Spandau UH, Rohrschneider K. Prevalence and geographical distribution of Usher syndrome in Germany. Graefes Arch Clin Exp Ophthalmol. 2002;240:495–8. doi: 10.1007/s00417-002-0485-8. Cohen M, Bitner-Glindzicz M, Luxon L. The changing face of Usher syndrome: clinical implications. Int J Audiol. 2007;46:82–93. doi: 10.1080/14992020600975279. Plantinga RF, Kleemola L, Huygen PL, Joensuu T, Sankila EM, Pennings RJ, Cremers CW. Serial audiometry and speech recognition findings in Finnish Usher syndrome type III patients. Audiol Neurootol. 2005;10:79–89. doi: 10.1159/000083363. Sadeghi M, Cohn ES, Kimberling WJ, Tranebjaerg L, Moller C. Audiological and vestibular features in affected subjects with USH3: a genotype/phenotype correlation. Int J Audiol. 2005;44:307–16. doi: 10.1080/14992020500060610. Pakarinen L, Tuppurainen K, Laippala P, Mantyjarvi M, Puhakka H. The ophthalmological course of Usher syndrome type III. Int Ophthalmol. 1995;19:307–11. doi: 10.1007/BF00130927. Plantinga RF, Pennings RJ, Huygen PL, Sankila EM, Tuppurainen K, Kleemola L, Cremers CW, Deutman AF. Visual impairment in Finnish Usher syndrome type III. Acta Ophthalmol Scand. 2006;84:36–41. doi: 10.1111/j.1600-0420.2005.00507. Herrera W, Aleman TS, Cideciyan AV, Roman AJ, Banin E, Ben-Yosef T, Gardner LM, Sumaroka A, Windsor EA, Schwartz SB, Stone EM, Liu XZ, Kimberling WJ, Jacobson SG. Retinal disease in Usher syndrome III caused by mutations in the clarin-1 gene. Invest Ophthalmol Vis Sci. 2008;49:2651–60. doi: 10.1167/iovs.07-1505. Weil D, Blanchard S, Kaplan J, Guilford P, Gibson F, Walsh J, Mburu P, Varela A, Levilliers J, Weston MD, Kelley PM, Kimberling WJ, Levi-Acobas MWF, Larget-Piet D, Munnich A, Steel KP, Brown SDM, Petit C. Defective myosin VIIA gene responsible for Usher syndrome type 1B. Nature. 1995;374:60–1. doi: 10.1038/374060a0. Bolz H, von Brederlow B, Ramírez A, Bryda EC, Kutsche K, Nothwang HG, Seeliger M. del C-Salcedó Cabrera M, Vila MC, Molina OP, Gal A, Kubisch C. Mutation of CDH23, encoding a new member of the cadherin gene family, causes Usher syndrome type 1D. Nat Genet. 2001;27:108–12. doi: 10.1038/83667. Ahmed ZM, Riazuddin S, Bernstein SL, Ahmed Z, Khan S, Griffith AJ, Morell RJ, Friedman TB, Riazuddin S, Wilcox ER. Mutations of the protocadherin gene PCDH15 cause Usher syndrome type 1F. Am J Hum Genet. 2001;69:25–34. doi: 10.1086/321277. Alagramam KN, Yuan H, Kuehn MH, Murcia CL, Wayne S, Srisailpathy CR, Lowry RB, Knaus R, Van Laer L, Bernier FP, Schwartz S, Lee C, Morton CC, Mullins RF, Ramesh A, Van Camp G, Hageman GS, Woychik RP, Smith RJ. Mutations in the novel protocadherin PCDH15 cause Usher syndrome type 1F. Hum Mol Genet. 2001;10:1709–18. doi: 10.1093/hmg/10.16.1709. Verpy E, Leibovici M, Zwaenepoel I, Liu XZ, Gal A, Salem N, Mansour A, Blanchard S, Kobayashi I, Keats BJ, Slim R, Petit C. A defect in harmonin, a PDZ domain-containing protein expressed in the inner ear sensory hair cells, underlies Usher syndrome type 1C. Nat Genet. 2000;26:51–5. doi: 10.1038/79171. Weil D, El-Amraoui A, Masmoudi S, Mustapha M, Kikkawa Y, Lainé S, Delmaghani S, Adato A, Nadifi S, Zina ZB, Hamel C, Gal A, Ayadi H, Yonekawa H, Petit C. Usher syndrome type I G (USH1G) is caused by mutations in the gene encoding SANS, a protein that associates with the USH1C protein, harmonin. Hum Mol Genet. 2003;12:463–71. doi: 10.1093/hmg/ddg051. Ebermann I, Scholl HP, Charbel Issa P, Becirovic E, Lamprecht J, Jurklies B, Millán JM, Aller E, Mitter D, Bolz H. A novel gene for Usher syndrome type 2: mutations in the long isoform of whirlin are associated with retinitis pigmentosa and sensorineural hearing loss. Hum Genet. 2007;121:203–11. doi: 10.1007/s00439-006-0304-0. Weston MD, Luijendijk MW, Humphrey KD, Moller C, Kimberling WJ. Mutations in the VLGR1 gene implicate G-protein signaling in the pathogenesis of Usher syndrome type II. Am J Hum Genet. 2004;74:357–66. doi: 10.1086/381685. Eudy JD, Yao S, Weston MD, Ma-Edmonds M, Talmadge CB, Cheng JJ, Kimberling WJ, Sumegi J. Isolation of a gene encoding a novel member of the nuclear receptor superfamily from the critical region of Usher syndrome type IIa at 1q41. Genomics. 1998;50:382–4. doi: 10.1006/geno.1998.5345. van Wijk E, Pennings RJ, te Brinke H, Claassen A, Yntema HG, Hoefsloot LH, Cremers FP, Cremers CW, Kremer H. Identification of 51 novel exons of the Usher syndrome type 2A (USH2A) gene that encode multiple conserved functional domains and that are mutated in patients with Usher syndrome type II. Am J Hum Genet. 2004;74:738–44. doi: 10.1086/383096. Joensuu T, Hämäläinen R, Yuan B, Johnson C, Tegelberg S, Gasparini P, Zelante L, Pirvola U, Pakarinen L, Lehesjoki AE, de la Chapelle A, Sankila EM. Mutations in a novel gene with transmembrane domains underlie Usher syndrome type 3. Am J Hum Genet. 2001;69:673–84. doi: 10.1086/323610. Adato A, Vreugde S, Joensuu T, Avidan N, Hamalainen R, Belenkiy O, Olender T, Bonne-Tamir B, Ben-Asher E, Espinos C, Millán JM, Lehesjoki AE, Flannery JG, Avraham KB, Pietrokovski S, Sankila EM, Beckmann JS, Lancet D. USH3A transcripts encode clarin-1, a four-transmembrane-domain protein with a possible role in sensory synapses. Eur J Hum Genet. 2002;10:339–50. doi: 10.1038/sj.ejhg.5200831. El-Amraoui A, Petit C. 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Hiroyuki Morimura , Gerald A. Fishman, Sandeep A. Grover, Anne B. Fulton, Eliot L. Berson, Thaddeus P. Dryja |   PNAS | 17 May 1998 | Vol. 95 (6) | pgs. 3088–3093 | doi:  10.1073/pnas.95.6.3088 Abstract RPE65 is a protein of unknown function expressed specifically by the retinal pigment epithelium. We examined all 14 exons of this gene in 147 unrelated patients with autosomal recessive retinitis pigmentosa (RP), in 15 patients with isolate RP, and in 45 patients with Leber congenital amaurosis (LCA). Sequence anomalies that were likely to be pathogenic were found in two patients with recessive RP, in one patient with isolate RP recategorized as recessive, and in seven patients with LCA. Cosegregation analysis in each available family showed that all affected individuals were either homozygotes or compound heterozygotes and that all unaffected individuals were either heterozygote carriers or homozygous wild type. In one family, there was one instance of a new mutation not present in either parent of the affected individual. In another family, affected members with recessive RP in three branches (i.e., three distinct pairs of parents) were compound heterozygotes for the same two mutations or homozygous for one of them. Based on our results, mutations in the RPE65 gene appear to account for ≈2% of cases of recessive RP and ≈16% of cases of LCA. Over 40 loci for human hereditary retinal degenerations involving the photoreceptors and retinal pigment epithelium are known ( http://utsph.sph.uth.tmc.edu/www/utsph/RetNet/home.htm ). Most of these loci are unidentified genes that have been recognized by linkage studies alone. Our group is engaged in identifying genes responsible for this set of diseases primarily through a candidate gene-based approach ( 1 ). In this report, we describe an analysis of the RPE65 gene, which has been assigned to chromosome 1p31 ( 2 ). This gene was discovered through its protein product that forms a complex with an antibody raised against human retinal pigment epithelium ( 3 ). The cDNA sequence predicts a protein with 533 amino acid residues and a molecular mass of ≈61 kDa ( 4 ). The protein is associated with the endoplasmic reticulum of the retinal pigment epithelium in vertebrates ( 3 , 5 ). Although the biochemical function of the protein product is currently obscure, the tissue-specific expression of the RPE65 gene made it an attractive candidate as a cause for some retinal degenerations because the retinal pigment epithelium has an essential role in maintaining the viability of the neighboring photoreceptor cells. In particular, enzymes in the endoplasmic reticulum are responsible for the recycling of the chromophore used by photoreceptor opsins. To explore the possible role that defects in RPE65 might play in the etiology of retinal degenerations, we screened a large cohort of unrelated patients with retinitis pigmentosa or Leber congenital amaurosis for mutations. Because RPE65 is expressed specifically in the eye, the study focused on patients with nonsyndromic forms of these diseases. This study was carried out contemporaneously with studies from two other groups that recently have been published ( 6 , 7 ) . Click here to read entire article References Dryja T P. Proc Natl Acad Sci USA. 1997;94:12117–12121. doi: 10.1073/pnas.94.22.12117. Hamel C P, Jenkins N A, Gilbert D J, Copeland N G, Redmond T M. Genomics. 1994;20:509–512. doi: 10.1006/geno.1994.1212. Hamel C P, Tsilou E, Harris E, Pfeffer B A, Hooks J J, Detrick B, Redmond T M. J Neurosci Res. 1993;34:414–425. doi: 10.1002/jnr.490340406. Hamel C P, Tsilou E, Pfeffer B A, Hooks J J, Detrick B, Redmond T M. J Biol Chem. 1993;268:15751–15757. Nicoletti A, Wong D J, Kawase K, Gibson L H, Yang-Feng T L, Richards J E, Thompson D A. Hum Mol Genet. 1995;4:641–649. doi: 10.1093/hmg/4.4.641. Marlhens F, Bareil C, Griffoin J M, Zrenner E, Amalric P, Eliaou C, Liu S Y, Harris E, Redmond T M, Arnaud B, et al. Nat Genet. 1997;17:139–141. doi: 10.1038/ng1097-139. Gu S M, Thompson D A, Srikumari C R S, Lorenz B, Finckh U, Nicoletti A, Murthy K R, Rathmann M, Kumaramanickavel G, Denton M J, et al. Nat Genet. 1997;17:194–197. doi: 10.1038/ng1097-194.

Mingchu Xu, Lizhu Yang, Feng Wang, Huajin Li,3 Xia Wang, Weichen Wang, Zhongqi Ge, Keqing Wang, Li Zhao, Hui Li, Yumei Li, Ruifang Sui, and Rui Chen | Human Genetics | 28 July 2015 | Vol 134 | 1069–1078  |  ncbi.nlm.nih.gov/pmc/articles/PMC4565766/ In this study, we totally investigated seven unrelated non-syndromic RD patients, including five RP and two LCA cases. Among them, five of them are Han Chinese and the remaining two are of European ethnicity diagnosed in United States. The index case we investigated, SRF71, is a 43-year-old male RP patient of Han Chinese ethnicity. . .  Preliminary screening by retinal capture sequencing found no causative mutations in known RP-causing genes. WES data show that he has biallelic variants in IFT140,  . . . Abstract Leber congenital amaurosis (LCA) and retinitis pigmentosa (RP) are two genetically heterogeneous retinal degenerative disorders. Despite the identification of a number of genes involved in LCA and RP, the genetic etiology remains unknown in many patients. In this study, we aimed to identify novel disease-causing genes of LCA and RP. Retinal capture sequencing was initially performed to screen mutations in known disease-causing genes in different cohorts of LCA and RP patients. For patients with negative results, we performed whole exome sequencing and applied a series of variant filtering strategies. Sanger sequencing was done to validate candidate causative IFT140 variants. Exome sequencing data analysis led to the identification of IFT140 variants in multiple unrelated non-syndromic LCA and RP cases. All the variants are extremely rare and predicted to be damaging. All the variants passed Sanger validation and segregation tests provided that the family members’ DNA was available. The results expand the phenotype spectrum of IFT140 mutations to non-syndromic retinal degeneration, thus extending our understanding of intraflagellar transport and primary cilia biology in the retina. This work also improves the molecular diagnosis of retinal degenerative disease. Introduction Leber congenital amaurosis (LCA, MIM# 204000) and retinitis pigmentosa (RP, MIM# 268000) are two types of inherited retinal degenerative diseases. LCA is featured by congenital or infantile-onset vision loss, nystagmus and absent electroretinogram (ERG) signals. RP is a more common and variable form of retinal degeneration (RD) and its onset ranges from childhood to mid-adulthood ( Hartong et al. 2006 ). Both LCA and RP are highly genetically heterogeneous. To date, at least 21 LCA-causing and 64 RP-causing genes have been identified (RetNet, the Retinal Information Network) (SP Daiger). Mutations in these genes account for about 70 % of LCA and 60 % of RP cases, respectively, suggesting that the molecular basis of a significant number of cases is yet to be discovered ( Wang et al. 2013 , 2014 ). Retinal degenerative disorders can be syndromic, in which case patients develop symptoms in other systems in addition to their ocular abnormalities. This phenomenon is frequently seen in ciliopathies with retinal involvement since photoreceptors develop highly specialized cilia structure and ciliated cells are widespread in the human body ( Hildebrandt et al. 2011 ). Mutations in ciliary genes were identified in a number of syndromes with RD including Senior–Løken syndrome (SLSN, MIM# 266900) ( Otto et al. 2005 ), Joubert syndrome (JBTS, MIM# 213300) ( Dixon-Salazar et al. 2004 ), Bardet–Biedl syndrome (BBS, MIM#209900) ( Mykytyn et al. 2001 ). It has also been reported that mutations in syndromic ciliopathy genes can lead to non-syndromic LCA or RP. For example, IQCB1 mutations were originally identified to cause SLSN ( Otto et al. 2005 ), but certain IQCB1 mutant alleles were found to cause LCA without renal symptoms ( Estrada-Cuzcano et al. 2011 ). Similarly, while mutations in CEP290 , a cilia basal body gene, can cause a series of syndromic ciliopathies including JBTS and BBS ( Baala et al. 2007 ; Sayer et al. 2006 ; Valente et al. 2006 ), it is also a major contributor to non-syndromic LCA cases ( den Hollander et al. 2006 ). Given the fact that cilia are responsible for numerous biological processes in multiple tissues, the diverse genotype–phenotype correlations observed by these studies can be explained by a combination of multi-functional nature of these ciliary genes and differential damaging effect of their mutant alleles. Intraflagellar transport (IFT) is a biological process by which various proteins are transported along the microtubule-based cilia ( Rosenbaum and Witman 2002 ). Specifically, the IFT-A complex is responsible for the return of proteins from the ciliary tip ( Absalon et al. 2008 ). Defects in IFT-A particles have already been associated with a spectrum of human ciliopathies. Mutations in one IFT-A complex component, WDR19 , were reported to cause cranioectodermal dysplasia (CED, MIM# 614378) ( Bredrup et al. 2011 ), nephronophthisis (NPHP, MIM# 614377) ( Halbritter et al. 2013b ) as well as non-syndromic retinitis pigmentosa ( Coussa et al. 2013 ). Similarly, mutations in IFT140 , another IFT-A complex gene, were known to cause two types of rare recessive ciliopathies: Mainzer–Saldino syndrome (MZSDS, MIM# 266920) and Jeune asphyxiating thoracic dystrophy (JATD, MIM# 208500) ( Khan et al. 2014 ; Perrault et al. 2012 ; Schmidts et al. 2013 ). MZSDS is featured by cone-shaped epiphysis, chronic renal disease, abnormality of the proximal femur and RD ( Beals and Weleber 2007 ; Giedion 1979 ). JATD patients show constricted thoracic cage, short-limbed short stature, polydactyly and often develop multi-organ disorders including retinal abnormalities ( Bard et al. 1978 ; de Vries et al. 2010 ; Oberklaid et al. 1977 ). Since both WDR19 and IFT140 are linked to ciliopathies with retinal involvement, it is intriguing for us to know, whether IFT140 defects, like WDR19 mutations, can also cause non-syndromic RD. In this study, through whole exome sequencing (WES), mutations in IFT140 have been identified in seven patients diagnosed with non-syndromic LCA or RP. These patients come from diverse ethnicities and account for about 1 % of non-syndromic RD cases. Our results highlight a novel genotype–phenotype correlation of a ciliary gene, which can improve the molecular diagnosis of retinal degenerative diseases and our understanding of intraflagellar transport in the retina. 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Melissa C. Humbert, Katie Weihbrecht, Charles C. Searby, Yalan Li, Robert M. Pope, Val C. Sheffield, Seongjin Seo | Proceedings of the National Academy of Sciences of the United States of America (PNAS) | 27 Nov 2012 | 109 (48) | 19691-19696 | doi.org/10.1073/pnas.1210916109 Abstract Mutations affecting ciliary components cause a series of related genetic disorders in humans, including nephronophthisis (NPHP), Joubert syndrome (JBTS), Meckel-Gruber syndrome (MKS), and Bardet-Biedl syndrome (BBS), which are collectively termed "ciliopathies." Recent protein-protein interaction studies combined with genetic analyses revealed that ciliopathy-related proteins form several functional networks/modules that build and maintain the primary cilium. However, the precise function of many ciliopathy-related proteins and the mechanisms by which these proteins are targeted to primary cilia are still not well understood. Here, we describe a protein-protein interaction network of inositol polyphosphate-5-phosphatase E (INPP5E), a prenylated protein associated with JBTS, and its ciliary targeting mechanisms. INPP5E is targeted to the primary cilium through a motif near the C terminus and prenyl-binding protein phosphodiesterase 6D (PDE6D) -dependent mechanisms. Ciliary targeting of INPP5E is facilitated by another JBTS protein, ADP-ribosylation factor-like 13B (ARL13B), but not by ARL2 or ARL3. ARL13B missense mutations that cause JBTS in humans disrupt the ARL13B-INPP5E interaction. We further demonstrate interactions of INPP5E with several ciliary and centrosomal proteins, including a recently identified ciliopathy protein centrosomal protein 164 (CEP164). These findings indicate that ARL13B, INPP5E, PDE6D, and CEP164 form a distinct functional network that is involved in JBTS and NPHP but independent of the ones previously defined by NPHP and MKS proteins. Primary cilia are microtubule-based cell surface projections that emanate from the centrosome. This subcellular organelle functions as an antenna, sensing and transducing extracellular signals into the cell, and plays an essential role in regulating multiple cellular processes including the cell cycle, embryonic development, and tissue homeostasis ( 1 – 3 ). Mutations affecting ciliary and centrosomal components underlie a group of related human disorders such as Joubert syndrome (JBTS), Meckel-Gruber syndrome (MKS), nephronophthisis (NPHP), and Bardet-Biedl syndrome (BBS), collectively termed ciliopathies ( 1 – 3 ). Recent protein–protein interaction studies have identified several functional modules or networks involved in these ciliopathies ( 4 ). For example, BBS proteins and intraflagellar transport (IFT) proteins form multiprotein complexes, the BBSome and the IFT complexes, respectively, and these complexes are involved in transporting ciliary proteins. Likewise, NPHP and MKS proteins form a distinct modular complex at the transition zone of primary cilia and regulate ciliary membrane compositions ( 5 – 9 ). However, there are many ciliary and centrosomal proteins [e.g., inositol polyphosphate-5-phosphatase E (INPP5E) and ADP-ribosylation factor-like 13B (ARL13B)] that have not been linked to any of the known functional networks and their precise functions remain to be elucidated. INPP5E encodes an enzyme that hydrolyzes the 5-phosphate of PtdIns(3,4,5)P3 and PtdIns(4,5)P2 and localizes to primary cilia. Mutations in this gene cause JBTS in humans ( 10 , 11 ). In mice, loss of Inpp5e activity results in cystic kidney, bilateral anophthalmia, polydactyly, skeletal defects, cleft palate, and cerebral developmental defects ( 11 ). Inactivation of Inpp5e in adult mice results in obesity and photoreceptor degeneration. Interestingly, many proteins that localize to cilia, including INPP5E, RPGR, PDE6 α and β subunits, GRK1 (Rhodopsin kinase), and GNGT1 (Transducin γ chain), are prenylated (either farnesylated or geranylgeranylated), and mutations in these genes or genes involved in their prenylation (e.g., AIPL1 and RCE1 ) lead to photoreceptor degeneration in vertebrates and humans ( 12 – 18 ). Understanding the molecular mechanisms by which prenylated proteins are transported to primary cilia would have significant ramifications in developing therapeutic strategies to treat blindness and other ciliopathy phenotypes associated with these genes. In this study, we sought to determine the functional network of INPP5E and the mechanisms by which INPP5E is targeted to primary cilia. We determined the ciliary targeting sequence (CTS) of INPP5E, the small GTPase responsible for its ciliary targeting, and an interaction network that connects INPP5E to other known ciliopathy genes and new candidates. 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