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Underlying genetic mechanisms of hereditary dystrophies in retina and cornea
Umeå University, Faculty of Medicine, Department of Medical Biosciences. (Irina Golovleva)ORCID iD: 0000-0002-8899-2520
2017 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Inherited retinal and corneal dystrophies represent a group of disorders with great genetic heterogeneity. Over 250 genes are associated with retinal diseases and 16 genes are causative of corneal dystrophies. This thesis is focused on finding the genetic causes of corneal dystrophy, Leber congenital amaurosis (LCA), Stargardt disease and retinitis pigmentosa in families from northern Sweden.  By whole exome sequencing a novel mutation, c.2816C>T, p.Thr939Ile, in Collagen Type XVII, Alpha 1 chain, COL17A1, gene was identified in several families with epithelial recurrent erosion dystrophy (ERED). We showed that the COL17A1 protein is expressed in the basement membrane of the cornea, explaining the mutation involvement in the corneal symptoms. We could link all the families in this study to a couple born in the late 1700s confirming a founder mutation in northern Sweden. Our finding highlights role of COL17A1 in ERED and suggests screening of this gene in patients with similar phenotype worldwide. Furthermore the genetic causes in several retinal degenerations were identified. In one family with two recessive disorders, LCA and Stargardt disease, a novel stop mutation, c.2557C>T, p.Gln853Stop, was detected in all LCA patients. In the Stargardt patients two intronic variants, the novel c.4773+3A>G and c.5461-10T>C, were detected in the ABCA4 gene. One individual was homozygous for the known variant c.5461-10T>C and the other one was compound heterozygote with both variants present. Both variants, c.4773+3A>G and c.5461-10T>C caused exon skipping in HEK293T cells demonstrated by in vitro splice assay, proving their pathogenicity in Stargardt disease. Finally, in recessive retinitis pigmentosa, Bothnia Dystrophy (BD), we identified a second mutation in the RLBP1 gene, c.677T>A, p.Met226Lys. Thus, BD is caused not only by common c.700C>T variant but also by homozygosity of c.677T>A or compound heterozygosity. Notably, known variant, c.40C>T, p.R14W in the CAIV gene associated with a dominant retinal dystrophy RP17 was detected in one of the compound BD heterozygote and his unaffected mother. This variant appears to be a benign variant in the population of northern Sweden.

In conclusion, novel genetic causes of retinal dystrophies in northern Sweden were found demonstrating the heterogeneity and complexity of retinal diseases. Identification of the genetic defect in COL17A1 in the corneal dystrophy contributes to understanding ERED pathogenesis and encourages refinement of IC3D classification. Our results provide valuable information for future molecular testing and genetic counselling of the families.

Place, publisher, year, edition, pages
Umeå: Umeå Universitet , 2017. , 57 p.
Series
Umeå University medical dissertations, ISSN 0346-6612 ; 1872
Keyword [en]
Cornea, retina, gene, mutation detection, inherited diseases
National Category
Genetics
Research subject
Genetics
Identifiers
URN: urn:nbn:se:umu:diva-130538ISBN: 978-91-7601-626-8 (print)OAI: oai:DiVA.org:umu-130538DiVA: diva2:1067797
Public defence
2017-02-17, Major Groove, Målpunkt J-11, Norrlands Universitetssjukhus, Umeå, 13:00 (English)
Opponent
Supervisors
Available from: 2017-01-27 Created: 2017-01-23 Last updated: 2017-02-01Bibliographically approved
List of papers
1. Mutations in Collagen, Type XVII, Alpha 1 (COL17A1) Cause Epithelial Recurrent Erosion Dystrophy (ERED)
Open this publication in new window or tab >>Mutations in Collagen, Type XVII, Alpha 1 (COL17A1) Cause Epithelial Recurrent Erosion Dystrophy (ERED)
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2015 (English)In: Human Mutation, ISSN 1059-7794, E-ISSN 1098-1004, Vol. 36, no 4, 463-473 p.Article in journal (Refereed) Published
Abstract [en]

Corneal dystrophies are a clinically and genetically heterogeneous group of inherited disorders that bilaterally affect corneal transparency. They are defined according to the corneal layer affected and by their genetic cause. In this study, we identified a dominantly inherited epithelial recurrent erosion dystrophy (ERED)-like disease that is common in northern Sweden. Whole-exome sequencing resulted in the identification of a novel mutation, c.2816C>T, p.T939I, in the COL17A1 gene, which encodes collagen type XVII alpha 1. The variant segregated with disease in a genealogically expanded pedigree dating back 200 years. We also investigated a unique COL17A1 synonymous variant, c.3156C>T, identified in a previously reported unrelated dominant ERED-like family linked to a locus on chromosome 10q23-q24 encompassing COL17A1. We show that this variant introduces a cryptic donor site resulting in aberrant pre-mRNA splicing and is highly likely to be pathogenic. Bi-allelic COL17A1 mutations have previously been associated with a recessive skin disorder, junctional epidermolysis bullosa, with recurrent corneal erosions being reported in some cases. Our findings implicate presumed gain-of-function COL17A1 mutations causing dominantly inherited ERED and improve understanding of the underlying pathology.

Place, publisher, year, edition, pages
John Wiley & Sons, 2015
Keyword
COL17A1, BP180, cornea dystrophy, ERED, ddPCR
National Category
Medical Bioscience
Identifiers
urn:nbn:se:umu:diva-103155 (URN)10.1002/humu.22764 (DOI)000352304200011 ()25676728 (PubMedID)
Note

Contract grant sponsors: Umeå University and Västerbotten County Council, Research and Development Foundation sponsored by Västerbotten County Council, Cronqvists Stiftelse (administered by The Swedish Society of Medicine); Ögonfonden, Stiftelsen KMA; the National Swedish Research Council (521-2013-2612); National Institute for Health Research Biomedical Research Centre at Moorfields Eye Hospital and UCL Institute of Ophthalmology; Moorfields Special Trustees; Moorfields Eye Charity; the Lanvern foundation.

Available from: 2015-05-29 Created: 2015-05-18 Last updated: 2017-01-23Bibliographically approved
2. Novel mutations in CRB1 and ABCA4 genes cause Leber congenital amaurosis and Stargardt disease in a Swedish family
Open this publication in new window or tab >>Novel mutations in CRB1 and ABCA4 genes cause Leber congenital amaurosis and Stargardt disease in a Swedish family
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2013 (English)In: European Journal of Human Genetics, ISSN 1018-4813, E-ISSN 1476-5438, Vol. 21, no 11, 1266-1271 p.Article in journal (Refereed) Published
Abstract [en]

This study aimed to identify genetic mechanisms underlying severe retinal degeneration in one large family from northern Sweden, members of which presented with early-onset autosomal recessive retinitis pigmentosa and juvenile macular dystrophy. The clinical records of affected family members were analysed retrospectively and ophthalmological and electrophysiological examinations were performed in selected cases. Mutation screening was initially performed with microarrays, interrogating known mutations in the genes associated with recessive retinitis pigmentosa, Leber congenital amaurosis and Stargardt disease. Searching for homozygous regions with putative causative disease genes was done by high-density SNP-array genotyping, followed by segregation analysis of the family members. Two distinct phenotypes of retinal dystrophy, Leber congenital amaurosis and Stargardt disease were present in the family. In the family, four patients with Leber congenital amaurosis were homozygous for a novel c.2557C>T (p.Q853X) mutation in the CRB1 gene, while of two cases with Stargardt disease, one was homozygous for c.5461-10T>C in the ABCA4 gene and another was carrier of the same mutation and a novel ABCA4 mutation c.4773+3A>G. Sequence analysis of the entire ABCA4 gene in patients with Stargardt disease revealed complex alleles with additional sequence variants, which were evaluated by bioinformatics tools. In conclusion, presence of different genetic mechanisms resulting in variable phenotype within the family is not rare and can challenge molecular geneticists, ophthalmologists and genetic counsellors.

Place, publisher, year, edition, pages
Nature Publishing Group, 2013
Keyword
CRB1, ABCA4, SNP-array, Stargardt disease, Leber congenital amaurosis
National Category
Medical Genetics
Identifiers
urn:nbn:se:umu:diva-82275 (URN)10.1038/ejhg.2013.23 (DOI)000325861500015 ()23443024 (PubMedID)
Available from: 2013-10-29 Created: 2013-10-29 Last updated: 2017-01-25Bibliographically approved
3. ABCA4 intronic variants c.4773+3A and c.5461-10T>C cause Stargardt disease due to defective splicing.: Intronic ABCA4 variants cause splice defects in Stargardt disease
Open this publication in new window or tab >>ABCA4 intronic variants c.4773+3A and c.5461-10T>C cause Stargardt disease due to defective splicing.: Intronic ABCA4 variants cause splice defects in Stargardt disease
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

Inherited retinal dystrophies (IRD) represent a group of progressive conditions affecting the retina. There is a great genetic heterogeneity causing IRD and to-date more than 250 genes are associated with autosomal dominant, autosomal recessive and X-linked IRD. Stargardt disease (#248200) or macular degeneration with flecks, type 1, STGD1 is associated with early onset, central visual impairment, frequent appearance of yellowish flecks and mutations in ABCA4 gene. In this study, two intronic variants potentially causing the Stargardt disease were functionally tested in HEK293T and ARPE-19 cells using in vitro splice minigene assay. The two variants, c.4773+3A>G and c.5461-10T>C in the ABCA4 gene encoding ATP binding cassette sub-family A, member 4 protein, responsible for transport of a vitamin A precursor in the photoreceptors of the retina, were present in two Stargardt patients, members of the same Swedish family. It was suggested that the disease in one of the patients was caused by homozygous variant c.5461-10T>C and by both variants, in the other patient. The c.5461–10T>C, the third most common variant in STGD1 patients always segregating with the disease was proposed to be a polymorphism, a risk allele and finally, a disease-associated mutation, though its functional impact remained unknown for a long time. Functional analysis of the ABCA4 variants are complicated due to predominant expression of the ABCA4  in photoreceptors, which means no affected tissue can be easily obtained from the patients.

This study provides the evidence that c.4773+3A>G and c.5461-10T>C cause aberrant splicing and are indeed disease-causative variants.

National Category
Genetics
Research subject
Genetics
Identifiers
urn:nbn:se:umu:diva-130535 (URN)
Available from: 2017-01-23 Created: 2017-01-23 Last updated: 2017-01-24
4. Carrier of R14W in carbonic anhydrase IV presents Bothnia dystrophy phenotype caused by two allelic mutations in RLBP1
Open this publication in new window or tab >>Carrier of R14W in carbonic anhydrase IV presents Bothnia dystrophy phenotype caused by two allelic mutations in RLBP1
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2008 (English)In: Investigative Ophthalmology and Visual Science, ISSN 0146-0404, E-ISSN 1552-5783, Vol. 49, no 7, 3172-3177 p.Article in journal (Refereed) Published
Abstract [en]

Purpose: Bothnia dystrophy (BD) is an autosomal recessive retinitis pigmentosa (arRP) associated with the c.700C>T mutation in the RLBP1 gene. Testing of patients with BD has revealed the c.700C>T mutation on one or both alleles. The purpose of this study was to elucidate the underlying genetic mechanisms along with a clinical evaluation of the heterozygous patients with BD.

Methods: Patients with BD heterozygous for the RLBP1 c.700C>T were tested for 848 mutations by arrayed primer-extension technology. Further mutation detection was performed by PCR-restriction fragment length polymorphism (RFLP), sequencing, denaturing (d)HLPC and allelic discrimination. The ophthalmic examinations were performed in all c.700C>T heterozygotes.

Results: The clinical findings in 10 BD heterozygotes were similar to those in the homozygotes. The presence of a second mutation, c.677T>A, corresponding to p.M226K was detected in all 10 cases. Segregation analysis showed that the mutations were allelic, and the patients were compound heterozygotes [c.677T>A]+[c.700C>T]. One of those patients was also a carrier of the c.40C>T corresponding to the p.R14W change in carbonic anhydrase IV (CAIV) associated with autosomal dominant RP, RP17. His mother, a carrier of the identical change was declared healthy after ophthalmic examination. This sequence variant was found in 6 of 143 tested blood donors.

Conclusions: The high frequency of arRP in northern Sweden is due to two mutations in the RLBP1 gene: c.677T>A and c.700C>T. BD is caused by the loss of CRALBP function due to changed physical features and impaired activity of retinoid binding. The CAIV p.R14W sequence variant found in one of the patients with a BD phenotype is a benign polymorphism in a population of northern Sweden.

Place, publisher, year, edition, pages
Association for Research in Vision and Ophthalmology, 2008
Keyword
Adolescent, Adult, Aged, Aged; 80 and over, Alleles, Amino Acid Substitution, Arginine, Carbonic Anhydrase IV/*genetics, Carrier Proteins/*genetics, Child, Cytosine, Female, Fundus Oculi, Genes; Recessive, Heterozygote, Homozygote, Humans, Male, Middle Aged, Mutation, Phenotype, Retinitis Pigmentosa/*genetics/pathology/physiopathology, Thymine, Tryptophan
National Category
Medical Genetics Ophthalmology
Research subject
Ophtalmology; Genetics
Identifiers
urn:nbn:se:umu:diva-26981 (URN)10.1167/iovs.07-1664 (DOI)000257124000051 ()
Available from: 2009-11-05 Created: 2009-11-05 Last updated: 2017-01-25Bibliographically approved

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