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Abstract Background Autosomal recessive non-syndromic hearing loss (NSHL) and cone dystrophies (CODs) are highly genetically and phenotypically heterogeneous disorders. In this study, we applied the whole exome sequencing (WES) to find the cause of HL and COD in an Iranian consanguineous family with three affected individuals. Methods Three members from an Iranian consanguineous family who were suffering from NSHL and visual impairment were ascertained in this study. Comprehensive clinical evaluations and genetic analysis followed by bioinformatic and co-segregation studies were performed to diagnose the cause of these phenotypes. Data were collected from 2020 to 2022. Results All cases showed congenital bilateral NSHL, decreased visual acuity, poor color discrimination, photophobia and macular atrophy. Moreover, cornea, iris and anterior vitreous were within normal limit in both eyes, decreased foveal sensitivity, central scotoma and generalized depression of visual field were seen in three cases. WES results showed two variants, a novel null variant (p.Trp548Ter) in the PDE6C gene causing COD type 4 (Achromatopsia) and a previously reported variant (p.Ile84Thr) in the PDZD7 gene causing NSHL. Both variants were found in the cis configuration on chromosome 10 with a genetic distance of about 8.3 cM, leading to their co-inheritance. However, two diseases could appear independently in subsequent generations due to crossover during meiosis. Conclusions Here, we could successfully determine the etiology of a seemingly complex phenotype in two adjacent genes. We identified a novel variant in the PDE6C gene, related to achromatopsia. Interestingly, this variant could cooperatively cause visual disorders: cone dystrophy and cone-rod dystrophy.

Key Clinical Message: The presence of more than one genetic/genomic disorder is not uncommon. It is therefore essential to continuously consider new signs and symptoms over time. Administration of gene therapy could be extremely difficult in particular situations. A 9‐month‐old boy presented to our department for evaluation of developmental delay. We found that he was affected by intermediate junctional epidermolysis bullosa (COL17A1, c.3766 + 1G > A, homozygous), Angelman syndrome (5,5 Mb deletion of 15q11.2‐q13.1), and autosomal recessive deafness type 57 (PDZD7, c.883C > T, homozygous). [ABSTRACT FROM AUTHOR]

Key Clinical Message The presence of more than one genetic/genomic disorder is not uncommon. It is therefore essential to continuously consider new signs and symptoms over time. Administration of gene therapy could be extremely difficult in particular situations. Abstract A 9‐month‐old boy presented to our department for evaluation of developmental delay. We found that he was affected by intermediate junctional epidermolysis bullosa (COL17A1, c.3766 + 1G > A, homozygous), Angelman syndrome (5,5 Mb deletion of 15q11.2‐q13.1), and autosomal recessive deafness type 57 (PDZD7, c.883C > T, homozygous).

Abstract Hearing loss is the most common sensory neural disorder in humans, and according to a WHO estimation, 5.5% (466 million) of people worldwide have disabling hearing loss. In this study, a Chinese family with prelingual sensorineural hearing loss was investigated. The affected individuals showed moderately severe hearing loss at all frequencies. Using target genome enrichment and high-throughput sequencing, the homozygous variant c.2372del; p.(Ser791fs) was identified in PDZD7. This variant lies in exon 15 of PDZD7 and results in a frame shift followed by an early stop codon. It is classified as pathogenic according to the ACMG/AMP guidelines and ClinGen specifications. Our study expands the pathogenic variant spectrum of PDZD7 and strengthens the clinical importance of this gene in patients with moderately severe hearing loss.

Hearing loss is the most common sensory neural disorder in humans, and according to a WHO estimation, 5.5% (466 million) of people worldwide have disabling hearing loss. In this study, a Chinese family with prelingual sensorineural hearing loss was investigated. The affected individuals showed moderately severe hearing loss at all frequencies. Using target genome enrichment and high-throughput sequencing, the homozygous variant c.2372del; p.(Ser791fs) was identified in PDZD7. This variant lies in exon 15 of PDZD7 and results in a frame shift followed by an early stop codon. It is classified as pathogenic according to the ACMG/AMP guidelines and ClinGen specifications. Our study expands the pathogenic variant spectrum of PDZD7 and strengthens the clinical importance of this gene in patients with moderately severe hearing loss. [ABSTRACT FROM AUTHOR]

Hearing relies on the transduction of sound-evoked vibrations into electrical signals, occurring in the stereocilia bundle of inner ear hair cells. The G protein-coupled receptor (GPCR) ADGRV1 and the multi-PDZ protein PDZD7 play a critical role in the formation and function of stereocilia through their scaffolding and signaling properties. During hair cell development, the GPCR activity of ADGRV1 is specifically inhibited by PDZD7 through an unknown mechanism. Here, we describe the key interactions mediated by the two N-terminal PDZ domains of PDZD7 and the cytoplasmic domain of ADGRV1. Both PDZ domains can bind to the C-terminal PDZ binding motif (PBM) of ADGRV1 with the critical contribution of atypical C-terminal β extensions. The two PDZ domains form a supramodule in solution, stabilized upon PBM binding. Interestingly, we showed that the stability and binding properties of the PDZ tandem are affected by two deafness-causing mutations located in the binding grooves of PDZD7 PDZ domains.

Abstract Background Retinitis Pigmentosa (RP) is a clinically and genetically heterogeneous disorder that results in inherited blindness. Despite the large number of genes identified, only ~ 60% of cases receive a genetic diagnosis using targeted-sequencing. The aim of this study was to design a whole genome sequencing (WGS) based approach to increase the diagnostic yield of complex Retinitis Pigmentosa cases. Methods WGS was conducted in three family members, belonging to one large apparent autosomal dominant RP family that remained unsolved by previous studies, using Illumina TruSeq library preparation kit and Illumina HiSeq X platform. Variant annotation, filtering and prioritization were performed using a number of open-access tools and public databases. Sanger sequencing of candidate variants was conducted in the extended family members. Results We have developed and optimized an algorithm, based on the combination of different open-access tools, for variant prioritization of WGS data which allowed us to reduce significantly the number of likely causative variants pending to be manually assessed and segregated. Following this algorithm, four heterozygous variants in one autosomal recessive gene (USH2A) were identified, segregating in pairs in the affected members. Additionally, two pathogenic alleles in ADGRV1 and PDZD7 could be contributing to the phenotype in one patient. Conclusions The optimization of a diagnostic algorithm for WGS data analysis, accompanied by a hypothesis-free approach, have allowed us to unmask the genetic cause of the disease in one large RP family, as well as to reassign its inheritance pattern which implies differences in the clinical management of these cases. These results contribute to increasing the number of cases with apparently dominant inheritance that carry causal mutations in recessive genes, as well as the possible involvement of various genes in the pathogenesis of RP in one patient. Moreover, our WGS-analysis approach, based on open-access tools, can easily be implemented by other researchers and clinicians to improve the diagnostic yield of additional patients with inherited retinal dystrophies.

Usher syndrome (USH) is the leading cause of hereditary hearing–vision loss in humans. PDZ domain-containing 7 (PDZD7) has been reported to be a modifier of and contributor to USH. PDZD7 co-localizes with USH2 proteins in the inner ear hair cells and is essential for ankle-link formation and stereocilia development. PDZD7 contains three PDZ domains and a low-complexity region between the last two PDZ domains, which has been overlooked in the previous studies. Here we characterized a well-folded harmonin homology domain (HHD) from the middle region and solved the PDZD7 HHD structure at the resolution of 1.49 Å. PDZD7 HHD adopts the same five-helix fold as other HHDs found in Harmonin and Whirlin; however, in PDZD7 HHD, a unique α1N helix occupies the canonical binding pocket, suggesting a distinct binding mode. Moreover, we found that the PDZD7 HHD domain can bind lipid and mediate the localization of PDZD7 to the plasma membrane in HEK293T cells. Intriguingly, a hearing-loss mutation at the N-terminal extension region of the HHD can disrupt the lipid-binding ability of PDZD7 HHD, suggesting that HHD-mediated membrane targeting is required for the hearing process. This structural and biochemical characterization of the PDZD7 HHD region provides mechanistic explanations for human deafness-causing mutations in PDZD7. Furthermore, this structure will also facilitate biochemical and functional studies of other HHDs.

Background: Retinitis Pigmentosa (RP) is a clinically and genetically heterogeneous disorder that results in inherited blindness. Despite the large number of genes identified, only ~ 60% of cases receive a genetic diagnosis using targeted-sequencing. The aim of this study was to design a whole genome sequencing (WGS) based approach to increase the diagnostic yield of complex Retinitis Pigmentosa cases.Methods: WGS was conducted in three family members, belonging to one large apparent autosomal dominant RP family that remained unsolved by previous studies, using Illumina TruSeq library preparation kit and Illumina HiSeq X platform. Variant annotation, filtering and prioritization were performed using a number of open-access tools and public databases. Sanger sequencing of candidate variants was conducted in the extended family members.Results: We have developed and optimized an algorithm, based on the combination of different open-access tools, for variant prioritization of WGS data which allowed us to reduce significantly the number of likely causative variants pending to be manually assessed and segregated. Following this algorithm, four heterozygous variants in one autosomal recessive gene (USH2A) were identified, segregating in pairs in the affected members. Additionally, two pathogenic alleles in ADGRV1 and PDZD7 could be contributing to the phenotype in one patient.Conclusions: The optimization of a diagnostic algorithm for WGS data analysis, accompanied by a hypothesis-free approach, have allowed us to unmask the genetic cause of the disease in one large RP family, as well as to reassign its inheritance pattern which implies differences in the clinical management of these cases. These results contribute to increasing the number of cases with apparently dominant inheritance that carry causal mutations in recessive genes, as well as the possible involvement of various genes in the pathogenesis of RP in one patient. Moreover, our WGS-analysis approach, based on open-access tools, can easily be implemented by other researchers and clinicians to improve the diagnostic yield of additional patients with inherited retinal dystrophies. [ABSTRACT FROM AUTHOR]

Autosomal recessive non-syndromic hearing loss (ARNSHL) is a highly heterogeneous genetic condition. PDZD7 has emerged as a new genetic etiology of ARNSHL. Biallelic mutations in the PDZD7 gene have been reported in two German families, four Iranian families, and a Pakistani family with ARNSHL. The effect of PDZD7 on ARNSHL in other population has yet to be elucidated. Two Chinese ARNSHL families, each of which had two affected siblings, were included in this study. The families underwent target region capture and high-throughput sequencing to analyze the exonic, splice-site, and intronic sequences of 128 genes. Furthermore, 1751 normal Chinese individuals served as controls, and 122 Chinese families segregating with apparent ARNSHL, who had been previously excluded for variants in the common deafness genes GJB2 and SLC26A4, were subjected to screening for candidate mutations. We identified a novel homozygous missense mutation (p.Arg66Leu) and novel compound heterozygous frameshift mutations (p.Arg56fsTer24 and p.His403fsTer36) in Chinese families with ARNSHL. This is the first report to identify PDZD7 as an ARNSHL-associated gene in the Chinese population. Our finding could expand the pathogenic spectrum and strengthens the clinical diagnostic role of the PDZD7 gene in ARNSHL patients. [ABSTRACT FROM AUTHOR]

Background: We describe molecular diagnosis in a complex consanguineous family: four offspring presented with combinations of three distinctive phenotypes; non-syndromic hearing loss (NSHL), an unusual skeletal phenotype comprising multiple fractures, cranial abnormalities and diaphyseal expansion, and significant developmental delay with microcephaly. We performed Chromosomal Microarray Analysis on the offspring with either the skeletal or developmental delay phenotypes, and linkage analysis and whole exome sequencing (WES) on all four children, parents and maternal aunt.Results: Chromosomal microarray and FISH analysis identified a de novo unbalanced translocation as a cause of the microcephaly and severe developmental delay. WES identified a NSHL-causing splice variant in an autosomal recessive deafness gene PDZD7 which resided in a linkage region and affected three of the children. In the two children diagnosed with an unusual skeletal phenotype, WES eventually disclosed a heterozygous COL1A1 variant which affects C-propetide cleavage site of COL1. The variant was inherited from an apparently unaffected mosaic father in an autosomal dominant fashion. After the discovery of the COL1A1 variant, the skeletal phenotype was diagnosed as a high bone mass form of osteogenesis imperfecta.Conclusions: Next generation sequencing offers an unbiased approach to molecular genetic diagnosis in highly heterogeneous and poorly characterised disorders and enables early diagnosis as well as detection of mosaicism. [ABSTRACT FROM AUTHOR]

Deafness is the most frequent sensory disorder. With over 90 genes and 110 loci causally implicated in non-syndromic hearing loss, it is phenotypically and genetically heterogeneous. Here, we investigate the genetic etiology of deafness in four families of Iranian origin segregating autosomal recessive non-syndromic hearing loss (ARNSHL). We used a combination of linkage analysis, homozygosity mapping, and a targeted genomic enrichment platform to simultaneously screen 90 known deafnesscausing genes for pathogenic variants. Variant segregation was confirmed by Sanger sequencing. Linkage analysis and homozygosity mapping showed segregation with the DFNB57 locus on chromosome 10 in two families. Targeted genomic enrichment with massively parallel sequencing identified causal variants in PDZD7: a homozygous missense variant (p.Gly103Arg) in one family and compound heterozygosity for missense (p. Met285Arg) and nonsense (p.Tyr500Ter) variants in the second family. Screening of two additional families identified two more variants: (p.Gly228Arg) and (p.Gln526Ter). Variant segregation with the hearing loss phenotype was confirmed in all families by Sanger sequencing. The missense variants are predicted to be deleterious, and the two nonsense mutations produce null alleles. This report is the first to show that mutations in PDZD7 cause ARNSHL, a finding that offers addition insight into the USH2 interactome. We also describe a novel likely disease-causing mutation in CIB2 and illustrate the complexity associated with gene identification in diseases that exhibit large genetic and phenotypic heterogeneity. [ABSTRACT FROM AUTHOR]

In cochlea, deafness-related protein PDZD7 is an indispensable component of the ankle link complex, which is critical for the maturation of inner-ear hair cell for sound perception. Ankle links, connecting the different rows of cochlear stereocilia, are essential for the staircase-like development of stereocilia. However, the molecular mechanism of how PDZD7 governs stereociliary development remains unknown. Here, we reported a novel PDZD7-binding partner, FCHSD2, identified by yeast two-hybrid screening. FCHSD2 was reported to be expressed in hair cell, where it co-operated with CDC42 and N-WASP to regulate the formation of cell protrusion. The association between FCHSD2 and PDZD7 was further confirmed in COS-7 cells. More importantly, we solved the complex structure of FCHSD2 tail with PDZD7 PDZ3 domain at 2.0 Å resolution. The crystal structure shows that PDZD7 PDZ3 adopts a typical PDZ domain topology, comprising five β strands and two α helixes. The PDZ-binding motif of FCHSD2 tail stretches through the αB/βB groove of PDZD7 PDZ3. Our study not only uncovers the interaction between FCHSD2 tail and PDZD7 PDZ3 at the atomic level, but also provides clues of connecting the ankle link complex with cytoskeleton dynamics for exploiting the molecular mechanism of stereociliary development., (© 2022 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.)

Hearing relies on the transduction of sound-evoked vibrations into electrical signals, occurring in the stereocilia bundle of inner ear hair cells. The G protein-coupled receptor (GPCR) ADGRV1 and the multi-PDZ protein PDZD7 play a critical role in the formation and function of stereocilia through their scaffolding and signaling properties. During hair cell development, the GPCR activity of ADGRV1 is specifically inhibited by PDZD7 through an unknown mechanism. Here, we describe the key interactions mediated by the two N-terminal PDZ domains of PDZD7 and the cytoplasmic domain of ADGRV1. Both PDZ domains can bind to the C-terminal PDZ binding motif (PBM) of ADGRV1 with the critical contribution of atypical C-terminal β extensions. The two PDZ domains form a supramodule in solution, stabilized upon PBM binding. Interestingly, we showed that the stability and binding properties of the PDZ tandem are affected by two deafness-causing mutations located in the binding grooves of PDZD7 PDZ domains., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Colcombet-Cazenave, Cordier, Zhu, Bouvier, Litsardaki, Laserre, Prevost, Raynal, Caillet-Saguy and Wolff.)

PDZD7, a PDZ domain-containing scaffold protein, is critical for the organization of Usher syndrome type 2 (USH2) interactome. Recently, biallelic PDZD7 variants have been associated with autosomal-recessive, non-syndromic hearing loss (ARNSHL). Indeed, we identified novel, likely pathogenic PDZD7 variants based on the American College of Medical Genetics and Genomics/Association for Molecular Pathology (ACMG/AMP) guidelines from Korean families manifesting putative moderate-to-severe prelingual ARNSHL; these were c.490C>T (p.Arg164Trp), c.1669delC (p.Arg557Glyfs*13), and c.1526G>A (p.Gly509Glu), with p.Arg164Trp being a predominantly recurring variant. Given the recurring missense variant (p.Arg164Trp) from our cohort, we compared the genotyping data using six short tandem-repeat (STR) markers within or flanking PDZD7 between four probands carrying p.Arg164Trp and 81 normal-hearing controls. We observed an identical haplotype across three out of six STR genotyping markers exclusively shared by two unrelated hearing impaired probands but not by any of the 81 normal-hearing controls, suggesting a potential founder effect. However, STR genotyping, based on six STR markers, revealed various p.Arg164Trp-linked haplotypes shared by all of the affected subjects. In conclusion, PDZD7 can be an important causative gene for moderate to severe ARNSHL in Koreans. Moreover, at least some, if not all, p.Arg164Trp alleles in Koreans could exert a potential founder effect and arise from diverse haplotypes as a mutational hot spot. [ABSTRACT FROM AUTHOR]

Usher syndrome (USH) is the leading genetic cause of combined hearing and vision loss. Among the three USH clinical types, type 2 (USH2) occurs most commonly. USH2A, GPR98, and WHRN are three known causative genes of USH2, whereas PDZD7 is a modifier gene found in USH2 patients. The proteins encoded by these four USH genes have been proposed to form a multiprotein complex, the USH2 complex, due to interactions found among some of these proteins in vitro, their colocalization in vivo, and mutual dependence of some of these proteins for their normal in vivo localizations. However, evidence showing the formation of the USH2 complex is missing, and details on how this complex is formed remain elusive. Here, we systematically investigated interactions among the intracellular regions of the four USH proteins using colocalization, yeast two-hybrid, and pull-down assays. We show that multiple domains of the four USH proteins interact among one another. Importantly, both WHRN and PDZD7 are required for the complex formation with USH2A and GPR98. In this USH2 quaternary complex, WHRN prefers to bind to USH2A, whereas PDZD7 prefers to bind to GPR98. Interaction between WHRN and PDZD7 is the bridge between USH2A and GPR98. Additionally, the USH2 quaternary complex has a variable stoichiometry. These findings suggest that a non-obligate, short term, and dynamic USH2 quaternary protein complex may exist in vivo. Our work provides valuable insight into the physiological role of the USH2 complex in vivo and informs possible reconstruction of the USH2 complex for future therapy., (© 2014 by The American Society for Biochemistry and Molecular Biology, Inc.)

The 10 different genes associated with the deaf/blind disorder, Usher syndrome, encode a number of structurally and functionally distinct proteins, most expressed as multiple isoforms/protein variants. Functional characterization of these proteins suggests a role in stereocilia development in cochlear hair cells, likely owing to adhesive interactions in hair bundles. In mature hair cells, homodimers of the Usher cadherins, cadherin 23 and protocadherin 15, interact to form a structural fiber, the tip link, and the linkages that anchor the taller stereocilia's actin cytoskeleton core to the shorter adjacent stereocilia and the elusive mechanotransduction channels, explaining the deafness phenotype when these molecular interactions are perturbed. The conundrum is that photoreceptors lack a synonymous mechanotransduction apparatus, and so a common theory for Usher protein function in the two neurosensory cell types affected in Usher syndrome is lacking. Recent evidence linking photoreceptor cell dysfunction in the shaker 1 mouse model for Usher syndrome to light-induced protein translocation defects, combined with localization of an Usher protein interactome at the periciliary region of the photoreceptors suggests Usher proteins might regulate protein trafficking between the inner and outer segments of photoreceptors. A distinct Usher protein complex is trafficked to the ribbon synapses of hair cells, and synaptic defects have been reported in Usher mutants in both hair cells and photoreceptors. This review aims to clarify what is known about Usher protein function at the synaptic and apical poles of hair cells and photoreceptors and the prospects for identifying a unifying pathobiological mechanism to explain deaf/blindness in Usher syndrome., (Copyright © 2013 Elsevier Ltd. All rights reserved.)