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Scelsi, Hailee F.; Barlow, Brett M.; Saccuzzo, Emily G.; Lieberman, Raquel L.
doi: 10.1002/humu.24238pmid: 34082484
Rare variants of the olfactomedin domain of myocilin are considered causative for inherited, early‐onset open‐angle glaucoma, with a misfolding toxic gain‐of‐function pathogenic mechanism detailed by 20 years of laboratory research. Myocilin variants are documented in the scientific literature and identified through large‐scale genetic sequencing projects such as those curated in the Genome Aggregation Database (gnomAD). In the absence of key clinical and laboratory information, however, the pathogenicity of any given variant is not clear, because glaucoma is a heterogeneous and prevalent age‐onset disease, and common variants are likely benign. In this review, we reevaluate the likelihood of pathogenicity for the ~100 nonsynonymous missense, insertion‐deletion, and premature termination of myocilin olfactomedin variants documented in the literature. We integrate available clinical, laboratory cellular, biochemical and biophysical data, the olfactomedin domain structure, and population genetics data from gnomAD. Of the variants inspected, ~50% can be binned based on a preponderance of data, leaving many of uncertain pathogenicity that motivate additional studies. Ultimately, the approach of combining metrics from different disciplines will likely resolve outstanding complexities regarding the role of this misfolding‐prone protein within the context of a multifactorial and prevalent ocular disease, and pave the way for new precision medicine therapeutics.
Shaukat, Irfan; Bakhos‐Douaihy, Dalal; Zhu, Yingying; Seaayfan, Elie; Demaretz, Sylvie; Frachon, Nadia; Weber, Stefanie; Kömhoff, Martin; Vargas‐Poussou, Rosa; Laghmani, Kamel
doi: 10.1002/humu.24217pmid: 33973684
Forrest, Iain S.; Chaudhary, Kumardeep; Vy, Ha My T.; Bafna, Shantanu; Kim, Soyeon; Won, Hong‐Hee; Loos, Ruth J.F.; Cho, Judy; Pasquale, Louis R.; Nadkarni, Girish N.; Rocheleau, Ghislain; Do, Ron
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Lopes‐Marques, Mónica; Pacheco, Ana Rita; Peixoto, Maria João; Cardoso, Ana Rita; Serrano, Catarina; Amorim, António; Prata, Maria João; Cooper, David N.; Azevedo, Luísa
doi: 10.1002/humu.24221pmid: 34015158
Understanding the role of common polymorphisms in modulating the clinical phenotype when they co‐occur with a disease‐causing lesion is of critical importance in medical genetics. We explored the impact of apparently neutral common polymorphisms, using the gene encoding the urea cycle enzyme, ornithine transcarbamylase (OTC), as a model system. Distinct combinations of genetic backgrounds embracing two missense polymorphisms were created in cis with the pathogenic p.Arg40His replacement. In vitro enzymatic assays revealed that the polymorphic variants were able to modulate OTC activity both in the presence or absence of the pathogenic lesion. First, we found that the combination of the minor alleles of polymorphisms p.Lys46Arg and p.Gln270Arg significantly enhanced enzymatic activity in the wild‐type protein. Second, enzymatic assays revealed that the minor allele of the p.Gln270Arg polymorphism was capable of ameliorating OTC activity when combined in cis with the pathogenic p.Arg40His replacement. Structural analysis predicted that the minor allele of the p.Gln270Arg polymorphism would serve to stabilize the OTC wild‐type protein, thereby corroborating the results of the experimental assays. Our findings demonstrate the potential importance of cis‐interactions between common polymorphic variants and pathogenic missense mutations and illustrate how standing genetic variation can modulate protein function.
Zhai, Yiwen; Zhang, Zhanhui; Shi, Panlai; Martin, Donna M.; Kong, Xiangdong
doi: 10.1002/humu.24222pmid: 34015165
Neurodevelopmental disorders (NDDs) are a genetically heterogeneous group of diseases, affecting 1%–3% of children. Whole‐exome sequencing (WES) has been widely used as a first‐tier tool for identifying genetic causes of rare diseases. Trio‐WES was performed in a cohort of 74 pedigrees with NDDs. Exome‐based copy number variant (CNV) calling was incorporated into the traditional single‐nucleotide variant (SNV) and small insertion/deletion (Indel) analysis pipeline for WES data. An overall positive diagnostic yield of 54.05% (40/74) was obtained in the pipeline of combinational SNV/Indel and CNV analysis, including 35.13% (26/74) from SNV/Indel analysis and 18.92% (14/74) from exome‐based CNV analysis, respectively. In total, SNV/Indel analysis identified 38 variants in 28 different genes, of which 24 variants were novel; exome‐based CNV analysis identified 14 CNVs, including 2 duplications and 12 deletions, which ranged from 440 bp (single exon) to 16.86 Mb (large fragment) in size. In particular, a hemizygous deletion of exon 1 in the SLC16A2 gene was detected. Based on the diagnostic results, two families underwent prenatal diagnosis and had unaffected babies. The incorporation of exome‐based CNV detection into conventional SNV/Indel analysis for a single trio‐WES test significantly improved the diagnostic rate, making WES a more powerful, practical, and cost‐effective tool in the clinical diagnosis of NDDs.
Upadhyai, Priyanka; Radhakrishnan, Periyasamy; Guleria, Vishal S.; Kausthubham, Neethukrishna; Nayak, Shalini S.; Superti‐Furga, Andrea; Girisha, Katta M.
doi: 10.1002/humu.24235pmid: 34057271
Biallelic loss of function variants in TRIP11 encoding for the Golgi microtubule‐associated protein 210 (GMAP‐210) causes the lethal chondrodysplasia achondrogenesis type 1A (ACG1A). Loss of TRIP11 activity has been shown to impair Golgi structure, vesicular transport, and results in loss of IFT20 anchorage to the Golgi that is vital for ciliary trafficking and ciliogenesis. Here, we report four fetuses, two each from two families, who were ascertained antenatally with ACG1A. Affected fetuses in both families are homozygous for the deep intronic TRIP11 variant, c.5457+81T>A, which was found in a shared region of homozygosity. This variant was found to cause aberrant transcript splicing and the retention of 77 base pairs of intron 18. The TRIP11 messenger RNA and protein levels were drastically reduced in fibroblast cells derived from one of the affected fetuses. Using immunofluorescence we also detected highly compacted Golgi apparatus in affected fibroblasts. Further, we observed a significant reduction in the frequency of ciliated cells and in the length of primary cilia in subject‐derived cell lines, not reported so far in patient cells with TRIP11 null or hypomorphic variants. Our findings illustrate how pathogenic variants in intronic regions of TRIP11 can impact transcript splicing, expression, and activity, resulting in ACG1A.
Rajeshwari, Mohan; Karthi, Sellamuthu; Singh, Reetu; Efthymiou, Stephanie; Gowda, Vykuntaraju K.; Varalakshmi, Perumal; Srinivasan, Varunvenkat M.; Houlden, Henry; Keller, Markus A.; Rizzo, William B.; Ashokkumar, Balasubramaniem
doi:
Pham, Duyen H.; Pitman, Melissa R.; Kumar, Raman; Jolly, Lachlan A.; Schulz, Renee; Gardner, Alison E.; Nys, Rebekah; Heron, Sarah E.; Corbett, Mark A.; Kothur, Kavitha; Gill, Deepak; Rajagopalan, Sulekha; Kolc, Kristy L.; Halliday, Benjamin J.; Robertson, Stephen P.;
Xing, Chao; Kanchwala, Mohammed; Rios, Jonathan J.; Hyatt, Tommy; Wang, Richard C.; Tran, An; Dougherty, Irene; Tovar‐Garza, Andrea; Purnadi, Christy; Kumar, Monique G.; Berk, David; Shinawi, Marwan; Irvine, Alan D.; Toledo‐Bahena, Mirna; Agim, Nnenna G.; Glass, Donald A.
Showing 1 to 10 of 12 Articles
Mutations in Na‐K‐2Cl co‐transporter, NKCC2, lead to type I Bartter syndrome (BS1), a life‐threatening kidney disease. Yet, our knowledge of the molecular regulation of NKCC2 mutants remains poor. Here, we aimed to identify the molecular pathogenic mechanisms of one novel and three previously reported missense NKCC2 mutations. Co‐immunolocalization studies revealed that all NKCC2 variants are not functional because they are not expressed at the cell surface due to retention in the endoplasmic reticulum (ER). Cycloheximide chase assays together with treatment by protein degradation and mannose trimming inhibitors demonstrated that the defect in NKCC2 maturation arises from ER retention and associated degradation (ERAD). Small interfering RNA (siRNA) knock‐down experiments revealed that the ER lectin OS9 is involved in the ERAD of NKCC2 mutants. 4‐phenyl butyric acid (4‐PBA) treatment mimicked OS9 knock‐down effect on NKCC2 mutants by stabilizing their immature forms. Importantly, out of the four studied mutants, only one showed an increased protein maturation upon treatment with glycerol. In summary, our study reveals that BS1 is among diseases linked to the ERAD pathway. Moreover, our data open the possibility that maturation of some ER retained NKCC2 variants is correctable by chemical chaperones offering, therefore, promising avenues in elucidating the molecular pathways governing the ERAD of NKCC2 folding mutants.
Biobanks with exomes linked to electronic health records (EHRs) enable the study of genetic pleiotropy between rare variants and seemingly disparate diseases. We performed robust clinical phenotyping of rare, putatively deleterious variants (loss‐of‐function [LoF] and deleterious missense variants) in ERCC6, a gene implicated in inherited retinal disease. We analyzed 213,084 exomes, along with a targeted set of retinal, cardiac, and immune phenotypes from two large‐scale EHR‐linked biobanks. In the primary analysis, a burden of deleterious variants in ERCC6 was strongly associated with (1) retinal disorders; (2) cardiac and electrocardiogram perturbations; and (3) immunodeficiency and decreased immunoglobulin levels. Meta‐analysis of results from the BioMe Biobank and UK Biobank showed a significant association of deleterious ERCC6 burden with retinal dystrophy (odds ratio [OR] = 2.6, 95% confidence interval [CI]: 1.5‐4.6; p = 8.7 × 10−4), atypical atrial flutter (OR = 3.5, 95% CI: 1.9‐6.5; p = 6.2 × 10−5), arrhythmia (OR = 1.5, 95% CI: 1.2–2.0; p = 2.7 × 10−3), and lymphocyte immunodeficiency (OR = 3.8, 95% CI: 2.1–6.8; p = 5.0 × 10−6). Carriers of ERCC6 LoF variants who lacked a diagnosis of these conditions exhibited increased symptoms, indicating underdiagnosis. These results reveal a unique genetic link among retinal, cardiac, and immune disorders and underscore the value of EHR‐linked biobanks in assessing the full clinical profile of carriers of rare variants.
Mutations in ALDH3A2 cause Sjögren–Larsson syndrome (SLS), a neuro‐ichthyotic condition due to the deficiency of fatty aldehyde dehydrogenase (FALDH). We screened for novel mutations causing SLS among Indian ethnicity, characterized the identified mutations in silico and in vitro, and retrospectively evaluated their role in phenotypic heterogeneity. Interestingly, asymmetric distribution of nonclassical traits was observed in our cases. Nerve conduction studies suggested intrinsic‐minus‐claw hands in two siblings, a novel neurological phenotype to SLS. Genetic testing revealed five novel homozygous ALDH3A2 mutations in six cases: Case‐1‐NM_000382.2:c.50C>A, NP_000373.1:p.(Ser17Ter); Case‐2‐NM_000382.2:c.199G>T, NP_000373.1:p.(Glu67Ter); Case‐3‐NM_000382.2:c.1208G>A, NP_000373.1:p.(Gly403Asp); Case‐4‐NM_000382.2:c.1325C>T, NP_000373.1:p.(Pro442Leu); Case‐5 and ‐6 NM_000382.2:c.1349G>A, NP_000373.1:p.(Trp450Ter). The mutations identified were predicted to be pathogenic and disrupt the functional domains of the FALDH. p.(Pro442Leu) at the C‐terminal α‐helix, might impair the substrate gating process. Mammalian expression studies with exon‐9 mutants confirmed the profound reduction in the enzyme activity. Diminished aldehyde‐oxidizing activity was observed with cases‐2 and 3. Cases‐2 and 3 showed epidermal hyperplasia with mild intracellular edema, spongiosis, hypergranulosis, and perivascular‐interstitial lymphocytic infiltrate and a leaky eosinophilic epidermis. The presence of keratin‐containing milia‐like lipid vacuoles implies defective lamellar secretion with p.(Gly403Asp). This study improves our understanding of the clinical and mutational diversity in SLS, which might help to fast‐track diagnostic and therapeutic interventions of this debilitating disorder.
doi: 10.1002/humu.24237pmid: 34082468
PCDH19 is a nonclustered protocadherin molecule involved in axon bundling, synapse function, and transcriptional coregulation. Pathogenic variants in PCDH19 cause infantile‐onset epilepsy known as PCDH19‐clustering epilepsy or PCDH19‐CE. Recent advances in DNA‐sequencing technologies have led to a significant increase in the number of reported PCDH19‐CE variants, many of uncertain significance. We aimed to determine the best approaches for assessing the disease relevance of missense variants in PCDH19. The application of the American College of Medical Genetics and Association for Molecular Pathology (ACMG‐AMP) guidelines was only 50% accurate. Using a training set of 322 known benign or pathogenic missense variants, we identified MutPred2, MutationAssessor, and GPP as the best performing in silico tools. We generated a protein structural model of the extracellular domain and assessed 24 missense variants. We also assessed 24 variants using an in vitro reporter assay. A combination of these tools was 93% accurate in assessing known pathogenic and benign PCDH19 variants. We increased the accuracy of the ACMG‐AMP classification of 45 PCDH19 variants from 50% to 94%, using these tools. In summary, we have developed a robust toolbox for the assessment of PCDH19 variant pathogenicity to improve the accuracy of PCDH19‐CE variant classification.
doi: 10.1002/humu.24239pmid: 34085356
CDAGS Syndrome is a rare congenital disorder characterized by Craniosynostosis, Delayed closure of the fontanelles, cranial defects, clavicular hypoplasia, Anal and Genitourinary malformations, and Skin manifestations. We performed whole exome and Sanger sequencing to identify the underlying molecular cause in five patients with CDAGS syndrome from four distinct families. Whole exome sequencing revealed biallelic rare variants that disrupt highly conserved nucleotides within the RNU12 gene. RNU12 encodes a small nuclear RNA that is a component of the minor spliceosome and is essential for minor intron splicing. Targeted sequencing confirmed allele segregation within the four families. All five patients shared the same rare mutation NC_000022.10:g.43011402C>T, which alters a highly conserved nucleotide within the precursor U12 snRNA 3ʹ extension. Each of them also carried a rare variant on the other allele that either disrupts the secondary structure or the Sm binding site of the RNU12 snRNA. Whole transcriptome sequencing analysis of lymphoblastoid cells identified 120 differentially expressed genes, and differential alternative splicing analysis indicated there was an enrichment of alternative splicing events in the patient. These findings provide evidence of the involvement of RNU12 in craniosynostosis, anal and genitourinary patterning, and cutaneous disease.