BBS1 branchpoint variant is associated with non-syndromic retinitis pigmentosa

BackgroundInherited retinal diseases (IRDs) can be caused by variants in >270 genes. The Bardet-Biedl syndrome 1 (BBS1) gene is one of these genes and may be associated with syndromic and non-syndromic autosomal recessive retinitis pigmentosa (RP). Here, we identified a branchpoint variant in BBS1 and assessed its pathogenicity by in vitro functional analysis.MethodsWhole genome sequencing was performed for three unrelated monoallelic BBS1 cases with non-syndromic RP. A fourth case received MGCM 105 gene panel analysis. Functional analysis using a midigene splice assay was performed for the putative pathogenic branchpoint variant in BBS1. After confirmation of its pathogenicity, patients were clinically re-evaluated, including assessment of non-ocular features of Bardet-Biedl syndrome.ResultsClinical assessments of probands showed that all individuals displayed non-syndromic RP with macular involvement. Through detailed variant analysis and prioritisation, two pathogenic variants in BBS1, the most common missense variant, c.1169T>G (p.(Met390Arg)), and a branchpoint variant, c.592-21A>T, were identified. Segregation analysis confirmed that in all families, probands were compound heterozygous for c.1169T>G and c.592-21A>T. Functional analysis of the branchpoint variant revealed a complex splicing defect including exon 8 and exon 7/8 skipping, and partial in-frame deletion of exon 8.ConclusionA putative severe branchpoint variant in BBS1, together with a mild missense variant, underlies non-syndromic RP in four unrelated individuals. To our knowledge, this is the first report of a pathogenic branchpoint variant in IRDs that results in a complex splice defect. In addition, this research highlights the importance of the analysis of non-coding regions in order to provide a conclusive molecular diagnosis.

Download Full-text

Novel LAT Pathogenic Variants in a POI Family and Its Role in the Ovary

Frontiers in Genetics ◽

10.3389/fgene.2021.764160 ◽

2021 ◽

Vol 12 ◽

Author(s):

Kun Chu ◽

Yi He ◽

Ziyuan Li ◽

Zhongxin Jiang ◽

Liang Wang ◽

...

Keyword(s):

Compound Heterozygous ◽

Ovarian Insufficiency ◽

Mutant Proteins ◽

Adverse Health Outcomes ◽

Pathogenic Variants ◽

Whole Exome ◽

Variant Analysis ◽

First Time

Premature ovarian insufficiency (POI) affects about 1% of women under 40 years and leads most often to definitive infertility with adverse health outcomes. Genetic factor has been reported to play an important role in POI. However, the genetic etiology remains unknown in the majority of the POI patients. Whole-exome sequencing and variant analysis were carried out in a POI pedigree. In vitro studies of the wild-type and mutant proteins were conducted in primary granulosa cells (GCs) and granulosa cell line. The result showed that the patients carried compound heterozygous nonsynonymous mutations (c.245C > T and c.181C > G) in LAT gene, which were identified to be transmitted from their parents. The two variants were assessed to affect residues that were conserved across different species examined, and were predicted to be deleterious by software predictions. Protein structure predicting result indicated that the two variants could alter their interactions with surrounding residues, which may change the internal structure of the LAT protein. Moreover, LAT protein expression in GCs was demonstrated for the first time, and further functional assays suggested that this mutation could reduce LAT expression and influence GC survival, which may contribute to the etiology of POI. In summary, we detect novel LAT pathogenic variants in a POI pedigree and report for the first time that LAT is present and functional in the GCs of the ovary. Our findings not only shed new light on the role of LAT in GCs, but also broaden the spectrum of genetic causes of POI.

Download Full-text

A combined in silico, in vitro and clinical approach to characterise novel pathogenic missense variants in PRPF31 in retinitis pigmentosa

10.1101/480343 ◽

2018 ◽

Author(s):

Gabrielle Wheway ◽

Liliya Nazlamova ◽

Nervine Meshad ◽

Samantha Hunt ◽

Nicola Jackson ◽

...

Keyword(s):

Retinitis Pigmentosa ◽

In Silico ◽

Autosomal Dominant ◽

Missense Variant ◽

Incomplete Penetrance ◽

Clinical Approach ◽

Loss Of Function ◽

Autosomal Dominant Retinitis Pigmentosa ◽

Missense Variants

AbstractAt least six different proteins of the spliceosome, including PRPF3, PRPF4, PRPF6, PRPF8, PRPF31 and SNRNP200, are mutated in autosomal dominant retinitis pigmentosa (adRP). These proteins have recently been shown to localise to the base of the connecting cilium of the retinal photoreceptor cells, elucidating this form of RP as a retinal ciliopathy. In the case of loss-of-function variants in these genes, pathogenicity can easily be ascribed. In the case of missense variants, this is more challenging. Furthermore, the exact molecular mechanism of disease in this form of RP remains poorly understood.In this paper we take advantage of the recently published cryo EM-resolved structure of the entire human spliceosome, to predict the effect of a novel missense variant in one component of the spliceosome; PRPF31, found in a patient attending the genetics eye clinic at Bristol Eye Hospital. Monoallelic variants in PRPF31 are a common cause of autosomal dominant retinitis pigmentosa (adRP) with incomplete penetrance. We use in vitro studies to confirm pathogenicity of this novel variant PRPF31 c.341T>A, p.Ile114Asn.This work demonstrates how in silico modelling of structural effects of missense variants on cryo-EM resolved protein complexes can contribute to predicting pathogenicity of novel variants, in combination with in vitro and clinical studies. It is currently a considerable challenge to assign pathogenic status to missense variants in these proteins.

Download Full-text

Clear evidence of LAMA5 gene biallelic truncating variants causing infantile nephrotic syndrome

Kidney360 ◽

10.34067/kid.0004952021 ◽

2021 ◽

pp. 10.34067/KID.0004952021

Author(s):

Yukimasa Taniguchi ◽

China Nagano ◽

Kiyotoshi Sekiguchi ◽

Atsushi Tashiro ◽

Noriko Sugawara ◽

...

Keyword(s):

Nephrotic Syndrome ◽

Splice Site ◽

Japanese Patients ◽

Compound Heterozygous ◽

Missense Variants ◽

Targeted Next Generation Sequencing ◽

Pathogenic Variants ◽

Infantile Nephrotic Syndrome ◽

Splice Site Variant

Background: Pathogenic variants in single genes encoding podocyte-associated proteins have been implicated in about 30% of steroid resistant nephrotic syndrome (SRNS) patients in children. However, LAMA5 gene biallelic variants have been identified in only 7 patients so far, and most are missense variants of unknown significance. Furthermore, no functional analysis had been conducted for all but one of these variants. Here, we report three patients with LAMA5 gene biallelic truncating variants manifesting infantile nephrotic syndrome and one SRNS case with biallelic LAMA5 missense variants. Methods: We conducted comprehensive gene screening of Japanese patients with severe proteinuria. Using targeted next-generation sequencing, 62 podocyte-related genes were screened in 407 unrelated patients with proteinuria. For the newly discovered LAMA5 variants, we conducted in vitro heterotrimer formation assays. Results: Biallelic truncating variants in the LAMA5 gene (NM_005560) were detected in 3 patients from 2 families. All patients presented with proteinuria within 6 months of age. Patients 1 and 2 were siblings possessing a nonsense variant (c.9232C>T, p.(Arg3078*)) and a splice site variant (c.1282+1G>A) that led to exon 9 skipping and a frameshift. Patient 3 had a remarkable irregular contour of the glomerular basement membrane. She was subsequently found to have a nonsense variant (c.8185C>T, p.(Arg2720*)) and the same splice site variant in patients 1 and 2. By in vitro heterotrimer formation assays, both truncating variants produced smaller laminin α5 proteins that nevertheless formed trimers with laminin β1 and γ1 chains. Patient 4 showed SRNS at the age of eight and carried compound heterozygous missense variants (c.1493C>T, p.(Ala498Val) and c.8399G>A, p.(Arg2800His)). Conclusions: Our patients showed clear evidence of biallelic LAMA5 truncating variants causing infantile nephrotic syndrome. We also discerned the clinical and pathological characteristics observed in LAMA5-related nephropathy. LAMA5 variant screening should be performed in congenital/infantile nephrotic syndrome patients.

Download Full-text

Biallelic ERBB3 loss-of-function variants are associated with a novel multisystem syndrome without congenital contracture

Orphanet Journal of Rare Diseases ◽

10.1186/s13023-019-1241-z ◽

2019 ◽

Vol 14 (1) ◽

Author(s):

Niu Li ◽

Yufei Xu ◽

Yi Zhang ◽

Guoqiang Li ◽

Tingting Yu ◽

...

Keyword(s):

Postnatal Growth ◽

Compound Heterozygous ◽

Loss Of Function ◽

Atrioventricular Canal ◽

Multiple Systems ◽

Feeding Difficulties ◽

Functional Studies ◽

Pathogenic Variants ◽

Compound Heterozygous Variants

Abstract Background Gain-of-function pathogenic variants of the Erb-B2 receptor tyrosine kinase 3 (ERBB3) gene contribute to the occurrence and development of a variety of human carcinomas through activation of phosphatidylinositol 3-kinase (PI3K)/AKT and extracellular signal-regulated kinase (ERK) signaling. ERBB3 gene homozygous germline variants, whose loss of function may cause autosomal recessive congenital contractural syndrome, were recently identified. This study aims to identify the disease-causing gene in a Chinese pedigree with variable phenotypes involving multiple systems, including developmental delay, postnatal growth retardation, transient lower limb asymmetry, facial malformations, atrioventricular canal malformation, bilateral nystagmus and amblyopia, feeding difficulties, immunodeficiency, anemia, and liver damage, but without congenital contracture. Methods Trio-whole exome sequencing (WES) was performed to identify the disease-causing gene in a 24-month-old Chinese female patient. The pathogenicity of the identified variants was evaluated using in silico tools and in vitro functional studies. Results Trio-WES revealed compound heterozygous variants of c.1253 T > C (p.I418T) and c.3182dupA (p.N1061Kfs*16) in the ERBB3 gene. Functional studies showed that p.I418T resulted in normal expression of ERBB3, which was capable of interacting with ERBB2. However, the variant impaired ERBB3 phosphorylation, consequently blocking ERBB2 phosphorylation and AKT and ERK activation. The truncated protein resulting from the c.3182dupA variant also lacked the capacity to activate downstream signaling pathways. Conclusions We report the first patient with a novel multisystem syndrome disorder without congenital contracture resulting from biallelic loss-of-function variants of ERBB3.

Download Full-text

Compound heterozygous inheritance of two novel COQ2 variants results in familial coenzyme Q deficiency

Orphanet Journal of Rare Diseases ◽

10.1186/s13023-020-01600-8 ◽

2020 ◽

Vol 15 (1) ◽

Author(s):

Aliaa H. Abdelhakim ◽

Avinash V. Dharmadhikari ◽

Sara D. Ragi ◽

Jose Ronaldo Lima de Carvalho ◽

Christine L. Xu ◽

...

Keyword(s):

Coenzyme Q ◽

Clinical Manifestations ◽

Missense Variant ◽

Neurological Involvement ◽

Cone Dystrophy ◽

Compound Heterozygous ◽

Pathogenic Variants ◽

Whole Exome ◽

Compound Heterozygous Variants ◽

End Stage

Abstract Background Primary coenzyme Q10 deficiency is a rare disease that results in diverse and variable clinical manifestations. Nephropathy, myopathy and neurologic involvement are commonly associated, however retinopathy has also been observed with certain pathogenic variants of genes in the coenzyme Q biosynthesis pathway. In this report, we describe a novel presentation of the disease that includes nephropathy and retinopathy without neurological involvement, and which is the result of a compound heterozygous state arising from the inheritance of two recessive potentially pathogenic variants, previously not described. Materials and methods Retrospective report, with complete ophthalmic examination, multimodal imaging, electroretinography, and whole exome sequencing performed on a family with three affected siblings. Results We show that affected individuals in the described family inherited two heterozygous variants of the COQ2 gene, resulting in a frameshift variant in one allele, and a predicted deleterious missense variant in the second allele (c.288dupC,p.(Ala97Argfs*56) and c.376C > G,p.(Arg126Gly) respectively). Electroretinography results were consistent with rod-cone dystrophy in the affected individuals. All affected individuals in the family exhibited the characteristic retinopathy as well as end-stage nephropathy, without evidence of any neurological involvement. Conclusions We identified two novel compound heterozygous variants of the COQ2 gene that result in primary coenzyme Q deficiency. Targeted sequencing of coenzyme Q biosynthetic pathway genes may be useful in diagnosing oculorenal clinical presentations syndromes not explained by more well known syndromes (e.g., Senior-Loken and Bardet-Biedl syndromes).

Download Full-text

Persistent Hypoglycemia with Polycystic Kidneys: A Rare Combination – A Case Report

Biomedicine Hub ◽

10.1159/000511389 ◽

2020 ◽

Vol 5 (3) ◽

pp. 1-6

Author(s):

Priya Prasher ◽

Katherine Redmond ◽

Hillarey Stone ◽

James Bailes ◽

Edward Nehus ◽

...

Keyword(s):

Case Report ◽

Promoter Region ◽

Molecular Genetic ◽

Missense Variant ◽

Compound Heterozygous ◽

Molecular Genetic Testing ◽

Coding Region ◽

Polycystic Kidney ◽

Homozygous State ◽

Pathogenic Variants

We present the case of an infant referred to our NICU born at 39 weeks’ gestation with persistent hypoglycemia with elevated insulin levels (HI) requiring diazoxide to maintain normoglycemia. Additionally, polycystic kidney disease (PKD) was detected by ultrasound. Molecular genetic testing revealed pathogenic variants in the <i>PMM2</i>gene, i.e., a variant in the promoter region and a missense variant in the coding region. The precoding variant was recently described in 11 European families with similar phenotypes, either in a homozygous state or as compound heterozygous with a pathogenic coding variant. In neonates with HI associated with PKD, this rare recessive disorder should be considered.

Download Full-text

Biallelic VPS35L pathogenic variants cause 3C/Ritscher-Schinzel-like syndrome through dysfunction of retriever complex

Journal of Medical Genetics ◽

10.1136/jmedgenet-2019-106213 ◽

2019 ◽

Vol 57 (4) ◽

pp. 245-253 ◽

Cited By ~ 3

Author(s):

Kohji Kato ◽

Yasuyoshi Oka ◽

Hideki Muramatsu ◽

Filipp F Vasilev ◽

Takanobu Otomo ◽

...

Keyword(s):

Core Protein ◽

Early Stage ◽

Critical Role ◽

Missense Variant ◽

Compound Heterozygous ◽

Chondrodysplasia Punctata ◽

Loss Of Function ◽

Skeletal Malformation ◽

Endosomal Membrane ◽

Pathogenic Variants

Background3C/Ritscher-Schinzel syndrome is characterised by congenital cranio-cerebello-cardiac dysplasia, where CCDC22 and WASHC5 are accepted as the causative genes. In combination with the retromer or retriever complex, these genes play a role in endosomal membrane protein recycling. We aimed to identify the gene abnormality responsible for the pathogenicity in siblings with a 3C/Ritscher-Schinzel-like syndrome, displaying cranio-cerebello-cardiac dysplasia, coloboma, microphthalmia, chondrodysplasia punctata and complicated skeletal malformation.MethodsExome sequencing was performed to identify pathogenic variants. Cellular biological analyses and generation of knockout mice were carried out to elucidate the gene function and pathophysiological significance of the identified variants.ResultsWe identified compound heterozygous pathogenic variants (c.1097dup; p.Cys366Trpfs*28 and c.2755G>A; p.Ala919Thr) in the VPS35L gene, which encodes a core protein of the retriever complex. The identified missense variant lacked the ability to form the retriever complex, and the frameshift variant induced non-sense-mediated mRNA decay, thereby confirming biallelic loss of function of VPS35L. In addition, VPS35L knockout cells showed decreased autophagic function in nutrient-rich and starvation conditions, as well as following treatment with Torin 1. We also generated Vps35l−/− mice and demonstrated that they were embryonic lethal at an early stage, between E7.5 and E10.5.ConclusionsOur results suggest that biallelic loss-of-function variants in VPS35L underlies 3C/Ritscher-Schinzel-like syndrome. Furthermore, VPS35L is necessary for autophagic function and essential for early embryonic development. The data presented here provide a new insight into the critical role of the retriever complex in fetal development.

Download Full-text

Ranking of non-coding pathogenic variants and putative essential regions of the human genome

Nature Communications ◽

10.1038/s41467-019-13212-3 ◽

2019 ◽

Vol 10 (1) ◽

Cited By ~ 13

Author(s):

Alex Wells ◽

David Heckerman ◽

Ali Torkamani ◽

Li Yin ◽

Jonathan Sebat ◽

...

Keyword(s):

Genome Organization ◽

Regulatory Elements ◽

Structural Features ◽

Loss Of Function ◽

3D Genome ◽

Coding Regions ◽

Pathogenic Variants ◽

Machine Learning Model ◽

Using Data

AbstractA gene is considered essential if loss of function results in loss of viability, fitness or in disease. This concept is well established for coding genes; however, non-coding regions are thought less likely to be determinants of critical functions. Here we train a machine learning model using functional, mutational and structural features, including new genome essentiality metrics, 3D genome organization and enhancer reporter data to identify deleterious variants in non-coding regions. We assess the model for functional correlates by using data from tiling-deletion-based and CRISPR interference screens of activity of cis-regulatory elements in over 3 Mb of genome sequence. Finally, we explore two user cases that involve indels and the disruption of enhancers associated with a developmental disease. We rank variants in the non-coding genome according to their predicted deleteriousness. The model prioritizes non-coding regions associated with regulation of important genes and with cell viability, an in vitro surrogate of essentiality.

Download Full-text

X-Linked Retinitis Pigmentosa Caused by Non-Canonical Splice Site Variants in RPGR

International Journal of Molecular Sciences ◽

10.3390/ijms22020850 ◽

2021 ◽

Vol 22 (2) ◽

pp. 850

Author(s):

Friederike Kortüm ◽

Sinja Kieninger ◽

Pascale Mazzola ◽

Susanne Kohl ◽

Bernd Wissinger ◽

...

Keyword(s):

Retinitis Pigmentosa ◽

Splice Site ◽

Exon Skipping ◽

Open Reading Frame ◽

Aberrant Splicing ◽

Reading Frame ◽

Pathogenic Variants ◽

Nucleotide Addition ◽

Intron 2

We aimed to validate the effect of non-canonical splice site variants in the RPGR gene in five patients from four families diagnosed with retinitis pigmentosa. Four variants located in intron 2 (c.154 + 3_154 + 6del), intron 3 (c.247 + 5G>A), intron 7 (c.779-5T>G), and intron 13 (c.1573-12A>G), respectively, were analyzed by means of in vitro splice assays. Splicing analysis revealed different aberrant splicing events, including exon skipping and intronic nucleotide addition, which are predicted to lead either to an in-frame deletion affecting relevant protein domains or to a frameshift of the open reading frame. Our data expand the landscape of pathogenic variants in RPGR, thereby increasing the genetic diagnostic rate in retinitis pigmentosa and allowing patients harboring the analyzed variants to be enrolled in clinical trials.

Download Full-text

Autosomal Recessive Retinitis Pigmentosa Associated with Three Novel REEP6 Variants in Chinese Population

Genes ◽

10.3390/genes12040537 ◽

2021 ◽

Vol 12 (4) ◽

pp. 537

Author(s):

Lujia Zhang ◽

Ya Li ◽

Litao Qin ◽

Yu Wu ◽

Bo Lei

Keyword(s):

Retinitis Pigmentosa ◽

Autosomal Recessive ◽

Cultured Cells ◽

Missense Variant ◽

Chinese Families ◽

Targeted Next Generation Sequencing ◽

Novel Variants ◽

Next Generation Sequencing Ngs ◽

Computational Predictions

Retinitis pigmentosa 77 is caused by mutations of REEP6 (MIM: 609346), which encodes a protein for the development of photoreceptors. Our study was to identify disease-causing variants in three Chinese families using targeted next-generation sequencing (NGS). Multiple lines of computational predictions combined with in vitro cellular experiments were applied to evaluate the pathogenicity of the newly found variants. Three novel variants in REEP6, including one missense variant, c.268G>C, one frameshift variant, c.468delC, and one splicing variant, c.598+1G>C, were found, while c.268G>C was detected in all probands. The three variants were classified as likely pathogenic by the American College of Medical Genetics and Genomics (ACMG). REEP6 variant proteins c.268G>C and c.468delC in cultured cells destabilized the REEP6 protein and induced intracellular inclusions. Our data suggested that REEP6 c.268G>C may be a recurrent causative variant in Chinese autosomal recessive retinitis pigmentosa patients.

Download Full-text