CFAP47 is a novel causative gene implicated in X-linked polycystic kidney disease.

Autosomal dominant polycystic kidney disease (ADPKD) is a well-described condition in which ~80% of cases have a genetic explanation, while the genetic basis of sporadic cystic kidney disease in adults remains unclear in ~30% of cases. This study aimed to identify novel genes associated with polycystic kidney disease (PKD) in patients with sporadic cystic kidney disease in which a clear genetic change was not identified in established genes. A next-generation sequencing panel analyzed known genes related to renal cysts in 118 sporadic cases, followed by whole-genome sequencing on 47 unrelated individuals without identified candidate variants. Three male patients were found to have rare missense variants in the X-linked gene Cilia And Flagella Associated Protein 47 (CFAP47). CFAP47 was expressed in primary cilia of human renal tubules, and knockout mice exhibited vacuolation of tubular cells and tubular dilation, providing evidence that CFAP47 is a causative gene involved in cyst formation. This discovery of CFAP47 as a newly identified gene associated with PKD, displaying X-linked inheritance, emphasizes the need for further cases to understand the role of CFAP47 in PKD.

Surgical Management of Iris Bombe in Muscle-Eye-Brain Disease.

A 2-year-old girl with severe muscular dystrophy presented with unilateral eye pain and corneal clouding. She was found to have absent red reflex, hypotonia, cerebral hypoplasia, and iris bombe on ultrasound biomicroscopy, a feature not previously reported in this syndrome. She responded favorably to surgical management. Iris bombe can be a cause of glaucoma in muscle-eye-brain disease. This highlights the importance of incorporating ultrasound biomicroscopy into the diagnostic algorithm of muscle-eye-brain disease and other types of congenital syndromic glaucoma. [J Pediatr Ophthalmol Strabismus. 2023;60(4):e35-e37.].

Variants in ACTC1 underlie distal arthrogryposis accompanied by congenital heart defects.

Contraction of the human sarcomere is the result of interactions between myosin cross-bridges and actin filaments. Pathogenic variants in genes such as MYH7, TPM1, and TNNI3 that encode parts of the cardiac sarcomere cause muscle diseases that affect the heart, such as dilated cardiomyopathy and hypertrophic cardiomyopathy. In contrast, pathogenic variants in homologous genes such as MYH2, TPM2, and TNNI2 that encode parts of the skeletal muscle sarcomere cause muscle diseases affecting skeletal muscle, such as distal arthrogryposis (DA) syndromes and skeletal myopathies. To date, there have been few reports of genes (e.g., MYH7) encoding sarcomeric proteins in which the same pathogenic variant affects skeletal and cardiac muscle. Moreover, none of the known genes underlying DA have been found to contain pathogenic variants that also cause cardiac abnormalities. We report five families with DA because of heterozygous missense variants in the gene actin, alpha, cardiac muscle 1 (ACTC1). ACTC1 encodes a highly conserved actin that binds to myosin in cardiac and skeletal muscle. Pathogenic variants in ACTC1 have been found previously to underlie atrial septal defect, dilated cardiomyopathy, hypertrophic cardiomyopathy, and left ventricular noncompaction. Our discovery delineates a new DA condition because of variants in ACTC1 and suggests that some functions of ACTC1 are shared in cardiac and skeletal muscle.

Biallelic mutations in LAMA5 disrupts a skeletal noncanonical focal adhesion pathway and produces a distinct bent bone dysplasia.

BACKGROUND: Beyond its structural role in the skeleton, the extracellular matrix (ECM), particularly basement membrane proteins, facilitates communication with intracellular signaling pathways and cell to cell interactions to control differentiation, proliferation, migration and survival. Alterations in extracellular proteins cause a number of skeletal disorders, yet the consequences of an abnormal ECM on cellular communication remains less well understood METHODS: Clinical and radiographic examinations defined the phenotype in this unappreciated bent bone skeletal disorder. Exome analysis identified the genetic alteration, confirmed by Sanger sequencing. Quantitative PCR, western blot analyses, immunohistochemistry, luciferase assay for WNT signaling were employed to determine RNA, proteins levels and localization, and dissect out the underlying cell signaling abnormalities.  Migration and wound healing assays examined cell migration properties. FINDINGS: This bent bone dysplasia resulted from biallelic mutations in LAMA5, the gene encoding the alpha-5 laminin basement membrane protein. This finding uncovered a mechanism of disease driven by ECM-cell interactions between alpha-5-containing laminins, and integrin-mediated focal adhesion signaling, particularly in cartilage. Loss of LAMA5 altered ß1 integrin signaling through the non-canonical kinase PYK2 and the skeletal enriched SRC kinase, FYN. Loss of LAMA5 negatively impacted the actin cytoskeleton, vinculin localization, and WNT signaling. INTERPRETATION: This newly described mechanism revealed a LAMA5-ß1 Integrin-PYK2-FYN focal adhesion complex that regulates skeletogenesis, impacted WNT signaling and, when dysregulated, produced a distinct skeletal disorder. FUNDING: Supported by NIH awards R01 AR066124, R01 DE019567, R01 HD070394, and U54HG006493, and Czech Republic grants INTER-ACTION LTAUSA19030, V18-08-00567 and GA19-20123S.

Mutations in MYLPF Cause a Novel Segmental Amyoplasia that Manifests as Distal Arthrogryposis.

We identified ten persons in six consanguineous families with distal arthrogryposis (DA) who had congenital contractures, scoliosis, and short stature. Exome sequencing revealed that each affected person was homozygous for one of two different rare variants (c.470G>T [p.Cys157Phe] or c.469T>C [p.Cys157Arg]) affecting the same residue of myosin light chain, phosphorylatable, fast skeletal muscle (MYLPF). In a seventh family, a c.487G>A (p.Gly163Ser) variant in MYLPF arose de novo in a father, who transmitted it to his son. In an eighth family comprised of seven individuals with dominantly inherited DA, a c.98C>T (p.Ala33Val) variant segregated in all four persons tested. Variants in MYLPF underlie both dominant and recessively inherited DA. Mylpf protein models suggest that the residues associated with dominant DA interact with myosin whereas the residues altered in families with recessive DA only indirectly impair this interaction. Pathological and histological exam of a foot amputated from an affected child revealed complete absence of skeletal muscle (i.e., segmental amyoplasia). To investigate the mechanism for this finding, we generated an animal model for partial MYLPF impairment by knocking out zebrafish mylpfa. The mylpfa mutant had reduced trunk contractile force and complete pectoral fin paralysis, demonstrating that mylpf impairment most severely affects limb movement. mylpfa mutant muscle weakness was most pronounced in an appendicular muscle and was explained by reduced myosin activity and fiber degeneration. Collectively, our findings demonstrate that partial loss of MYLPF function can lead to congenital contractures, likely as a result of degeneration of skeletal muscle in the distal limb.

An alternative technique for pituitary tumour removal: Sequential endoscopic-microscopic pituitary procedure (SEMPP).

Several authors have reported advantages of the purely endoscopic (PE) approach over traditional purely microscopic (PM) techniques for many sinus and anterior skull base procedures. However, in pituitary surgery, the PE approach carries a number of limitations. Experienced microscopically-trained neurosurgeons are required to master the endoscopic technique which has a steep learning curve due to its one-handed nature and optically-distorted 2-dimensional view. We describe our novel technique, the sequential endoscopic and microscopic pituitary procedure (SEMPP) which does not require microscopically-trained neurosurgeons to alter their technique. We compare SEMPP with the PE approach in terms of outcome and safety. Retrospective chart review of consecutive SEMPP cases performed at our institution between January 2010 and December 2013 was conducted. Operative time, gross total resection rate, resolution of endocrine and visual dysfunction, hospital length of stay, cerebrospinal fluid (CSF) leak rate and revision rates were recorded. 32 patients were identified (50% female, mean age 53.0 years), and 33 SEMPP cases. Mean operating time was 132 min (range 90-200). 69% of patients experienced gross total resection. Most patients (81.3%) with preoperative visual deficit either experienced complete resolution or improved symptoms. The remainder experienced no change in vision. 12.5% (n = 4) of patients experienced intraoperative CSF leaks. All were repaired intraoperatively or with conservative management. Two patients (6.3%) experienced epistaxis managed with conservative measures. The SEMPP technique demonstrates comparable outcomes, complication rates and operative time to PE and PM techniques described in the literature.

A presynaptic congenital myasthenic syndrome attributed to a homozygous sequence variant in LAMA5.

We report a severe defect of neuromuscular transmission in a consanguineous patient with a homozygous variant in the laminin α5 subunit gene (LAMA5). The variant c.8046C > T (p.Arg2659Trp) is rare and has a predicted deleterious effect. The affected individual, who also carries a rare homozygous sequence variant in LAMA1, had normal cognitive function, but magnetic resonance brain imaging showed mild volume loss and periventricular T2 prolongation. Repetitive nerve stimulation at 2 Hz showed 50% decrement of compound muscle action potential amplitudes but 250% facilitation immediately after exercise, similar to that seen in Lambert-Eaton myasthenic syndrome. Endplate studies demonstrated a profound reduction of the endplate potential quantal content but normal amplitudes of miniature endplate potentials. Electron microscopy showed endplates with increased postsynaptic folding that were denuded or only partially occupied by small nerve terminals. Expression studies revealed that p.Arg2659Trp caused decreased binding of laminin α5 to SV2A and impaired laminin-521 cell adhesion and cell projection support in primary neuronal cultures. In summary, this report describing severe neuromuscular transmission failure in a patient with a LAMA5 mutation expands the list of phenotypes associated with defects in genes encoding α-laminins.

Variation in Cilia Protein Genes and Progression of Lung Disease in Cystic Fibrosis.

RATIONALE: Cystic fibrosis, like primary ciliary dyskinesia, is an autosomal recessive disorder characterized by abnormal mucociliary clearance and obstructive lung disease. We hypothesized that genes underlying the development or function of cilia may modify lung disease severity in persons with cystic fibrosis. OBJECTIVES: To test this hypothesis, we compared variants in 93 candidate genes in both upper and lower tertiles of lung function in a large cohort of children and adults with cystic fibrosis with those of a population control dataset. METHODS: Variants within candidate genes were tested for association using the SKAT-O test, comparing cystic fibrosis cases defined by poor (n = 127) or preserved (n = 127) lung function with population controls (n = 3,269 or 3,148, respectively). Associated variants were then tested for association with related phenotypes in independent datasets. RESULTS: Variants in DNAH14 and DNAAF3 were associated with poor lung function in cystic fibrosis, whereas variants in DNAH14 and DNAH6 were associated with preserved lung function in cystic fibrosis. Associations between DNAH14 and lung function were replicated in disease-related phenotypes characterized by obstructive lung disease in adults. CONCLUSIONS: Genetic variants within DNAH6, DNAH14, and DNAAF3 are associated with variation in lung function among persons with cystic fibrosis.

Genetic analysis of CHARGE syndrome identifies overlapping molecular biology.

PURPOSE: CHARGE syndrome is an autosomal-dominant, multiple congenital anomaly condition characterized by vision and hearing loss, congenital heart disease, and malformations of craniofacial and other structures. Pathogenic variants in CHD7, encoding adenosine triphosphate-dependent chromodomain helicase DNA binding protein 7, are present in the majority of affected individuals. However, no causal variant can be found in 5-30% (depending on the cohort) of individuals with a clinical diagnosis of CHARGE syndrome. METHODS: We performed whole-exome sequencing (WES) on 28 families from which at least one individual presented with features highly suggestive of CHARGE syndrome. RESULTS: Pathogenic variants in CHD7 were present in 15 of 28 individuals (53.6%), whereas 4 (14.3%) individuals had pathogenic variants in other genes (RERE, KMT2D, EP300, or PUF60). A variant of uncertain clinical significance in KDM6A was identified in one (3.5%) individual. The remaining eight (28.6%) individuals were not found to have pathogenic variants by WES. CONCLUSION: These results demonstrate that the phenotypic features of CHARGE syndrome overlap with multiple other rare single-gene syndromes. Additionally, they implicate a shared molecular pathology that disrupts epigenetic regulation of multiple-organ development.

ERCC4 variants identified in a cohort of patients with segmental progeroid syndromes.

Pathogenic variants in genes, which encode DNA repair and damage response proteins, result in a number of genomic instability syndromes with features of accelerated aging. ERCC4 (XPF) encodes a protein that forms a complex with ERCC1 and is required for the 5' incision during nucleotide excision repair. ERCC4 is also FANCQ, illustrating a critical role in interstrand crosslink repair. Pathogenic variants in this gene cause xeroderma pigmentosum, XFE progeroid syndrome, Cockayne syndrome (CS), and Fanconi anemia. We performed massive parallel sequencing for 42 unsolved cases submitted to the International Registry of Werner Syndrome. Two cases, each carrying two novel heterozygous ERCC4 variants, were identified. The first case was a compound heterozygote for: c.2395C > T (p.Arg799Trp) and c.388+1164_792+795del (p.Gly130Aspfs*18). Further molecular and cellular studies indicated that the ERCC4 variants in this patient are responsible for a phenotype consistent with a variant of CS. The second case was heterozygous for two variants in cis: c.[1488A > T; c.2579C > A] (p.[Gln496His; Ala860Asp]). While the second case also had several phenotypic features of accelerated aging, we were unable to provide biological evidence supporting the pathogenic roles of the associated ERCC4 variants. Precise genetic causes and disease mechanism of the second case remains to be determined.

A postnatal role for embryonic myosin revealed by MYH3 mutations that alter TGFß signaling and cause autosomal dominant spondylocarpotarsal synostosis.

Spondylocarpotarsal synostosis (SCT) is a skeletal disorder characterized by progressive vertebral, carpal and tarsal fusions, and mild short stature. The majority of affected individuals have an autosomal recessive form of SCT and are homozygous or compound heterozygous for nonsense mutations in the gene that encodes the cytoskeletal protein filamin B (FLNB), but a subset do not have FLNB mutations. Exome sequence analysis of three SCT patients negative for FLNB mutations identified an autosomal dominant form of the disease due to heterozygosity for missense or nonsense mutations in MYH3, which encodes embryonic myosin. Cells transfected with the MYH3 missense mutations had reduced TGFß signaling, revealing a regulatory role for embryonic myosin in the TGFß signaling pathway. In wild-type mice, there was persistent postnatal expression of embryonic myosin in the small muscles joining the neural arches of the spine suggesting that loss of myosin function in these muscles contribute to the disease. Our findings demonstrate that dominant mutations in MYH3 underlie autosomal dominant SCT, identify a postnatal role for embryonic myosin and suggest that altered regulation of signal transduction in the muscles within the spine may lead to the development of vertebral fusions.

Exome Sequencing of Phenotypic Extremes Identifies CAV2 and TMC6 as Interacting Modifiers of Chronic Pseudomonas aeruginosa Infection in Cystic Fibrosis.

Discovery of rare or low frequency variants in exome or genome data that are associated with complex traits often will require use of very large sample sizes to achieve adequate statistical power. For a fixed sample size, sequencing of individuals sampled from the tails of a phenotype distribution (i.e., extreme phenotypes design) maximizes power and this approach was recently validated empirically with the discovery of variants in DCTN4 that influence the natural history of P. aeruginosa airway infection in persons with cystic fibrosis (CF; MIM219700). The increasing availability of large exome/genome sequence datasets that serve as proxies for population-based controls affords the opportunity to test an alternative, potentially more powerful and generalizable strategy, in which the frequency of rare variants in a single extreme phenotypic group is compared to a control group (i.e., extreme phenotype vs. control population design). As proof-of-principle, we applied this approach to search for variants associated with risk for age-of-onset of chronic P. aeruginosa airway infection among individuals with CF and identified variants in CAV2 and TMC6 that were significantly associated with group status. These results were validated using a large, prospective, longitudinal CF cohort and confirmed a significant association of a variant in CAV2 with increased age-of-onset of P. aeruginosa airway infection (hazard ratio = 0.48, 95% CI=[0.32, 0.88]) and variants in TMC6 with diminished age-of-onset of P. aeruginosa airway infection (HR = 5.4, 95% CI=[2.2, 13.5]) A strong interaction between CAV2 and TMC6 variants was observed (HR=12.1, 95% CI=[3.8, 39]) for children with the deleterious TMC6 variant and without the CAV2 protective variant. Neither gene showed a significant association using an extreme phenotypes design, and conditions for which the power of an extreme phenotype vs. control population design was greater than that for the extreme phenotypes design were explored.

Autosomal-Dominant Multiple Pterygium Syndrome Is Caused by Mutations in MYH3.

Multiple pterygium syndrome (MPS) is a phenotypically and genetically heterogeneous group of rare Mendelian conditions characterized by multiple pterygia, scoliosis, and congenital contractures of the limbs. MPS typically segregates as an autosomal-recessive disorder, but rare instances of autosomal-dominant transmission have been reported. Whereas several mutations causing recessive MPS have been identified, the genetic basis of dominant MPS remains unknown. We identified four families affected by dominantly transmitted MPS characterized by pterygia, camptodactyly of the hands, vertebral fusions, and scoliosis. Exome sequencing identified predicted protein-altering mutations in embryonic myosin heavy chain (MYH3) in three families. MYH3 mutations underlie distal arthrogryposis types 1, 2A, and 2B, but all mutations reported to date occur in the head and neck domains. In contrast, two of the mutations found to cause MPS in this study occurred in the tail domain. The phenotypic overlap among persons with MPS, coupled with physical findings distinct from other conditions caused by mutations in MYH3, suggests that the developmental mechanism underlying MPS differs from that of other conditions and/or that certain functions of embryonic myosin might be perturbed by disruption of specific residues and/or domains. Moreover, the vertebral fusions in persons with MPS, coupled with evidence of MYH3 expression in bone, suggest that embryonic myosin plays a role in skeletal development.

De novo mutations in NALCN cause a syndrome characterized by congenital contractures of the limbs and face, hypotonia, and developmental delay.

Freeman-Sheldon syndrome, or distal arthrogryposis type 2A (DA2A), is an autosomal-dominant condition caused by mutations in MYH3 and characterized by multiple congenital contractures of the face and limbs and normal cognitive development. We identified a subset of five individuals who had been putatively diagnosed with "DA2A with severe neurological abnormalities" and for whom congenital contractures of the limbs and face, hypotonia, and global developmental delay had resulted in early death in three cases; this is a unique condition that we now refer to as CLIFAHDD syndrome. Exome sequencing identified missense mutations in the sodium leak channel, non-selective (NALCN) in four families affected by CLIFAHDD syndrome. We used molecular-inversion probes to screen for NALCN in a cohort of 202 distal arthrogryposis (DA)-affected individuals as well as concurrent exome sequencing of six other DA-affected individuals, thus revealing NALCN mutations in ten additional families with "atypical" forms of DA. All 14 mutations were missense variants predicted to alter amino acid residues in or near the S5 and S6 pore-forming segments of NALCN, highlighting the functional importance of these segments. In vitro functional studies demonstrated that NALCN alterations nearly abolished the expression of wild-type NALCN, suggesting that alterations that cause CLIFAHDD syndrome have a dominant-negative effect. In contrast, homozygosity for mutations in other regions of NALCN has been reported in three families affected by an autosomal-recessive condition characterized mainly by hypotonia and severe intellectual disability. Accordingly, mutations in NALCN can cause either a recessive or dominant condition characterized by varied though overlapping phenotypic features, perhaps based on the type of mutation and affected protein domain(s).

Schneiderian-Type Papilloma of the Middle Ear: A Review of the Literature.

Schneiderian-type papilloma of the middle ear is a rare finding. We present a 46-year-old Aboriginal man with a large tympanic membrane perforation and a Schneiderian-type papilloma filling the middle ear. The aim of this study is to familiarize clinicians with this uncommon disease through discussion of its clinical presentation, diagnostic considerations and management. A search of English-language peer-reviewed literature was undertaken using the key words "Schneiderian-type papilloma," "inverted papilloma," and "middle ear." A total of 29 cases (including the present case) of Schneiderian-type papilloma involving the middle ear were reviewed. Common presenting symptoms include hearing loss, otalgia, and otorrhea. Middle ear disease is associated with higher rates of recurrence and malignant transformation than its sinonasal counterpart. Radical surgical resection is the only curative treatment. Schneiderian-type papilloma is a benign, but locally aggressive, epithelial neoplasm most commonly arising in the sinonasal tract. Whilst involvement of the middle ear is extremely rare, knowledge of this condition is important due to its propensity to recur and the high rate of malignant transformation.

Disclosure of genetic research results to members of a founder population.

There is currently extensive discussion and debate in the literature on how, when, and to whom genetic research results should be returned (see Genetics in Medicine, April 2012 issue). Here, we describe our experience in disclosing genetic information on Mendelian disorders discovered during the course of our research in the Hutterites. We first assessed attitudes toward the disclosure of carrier results, which revealed that many individuals wanted carrier information and that many intended to use the information in family planning. Based on this information, we developed a pilot study to test and disclose cystic fibrosis (CF) carrier status. Next, a larger scale project was developed in order to disclose genetic research results for 14 diseases to those interested in receiving the information. We developed brochures, offered a live interactive educational program, conducted a consent process, and disclosed results in letters mailed to the consented individuals. Overall, ~80% of individuals who participated in the educational program signed consent forms for the release of their results for 14 diseases. We describe our experience with returning individual genetic research results to participants in a population-based research study.

Homozygous founder mutation in desmocollin-2 (DSC2) causes arrhythmogenic cardiomyopathy in the Hutterite population.

BACKGROUND: Dominant mutations in cellular junction proteins are the major cause of arrhythmogenic cardiomyopathy, whereas recessive mutations in those proteins cause cardiocutaneous syndromes such as Naxos and Carvajal syndrome. The Hutterites are distinct genetic isolates who settled in North America in 1874. Descended from <100 founders, they trace their origins to 16th-century Europe. METHODS AND RESULTS: We clinically and genetically evaluated 2 large families of the Alberta Hutterite population with a history of sudden death and found several individuals with severe forms of biventricular cardiomyopathy characterized by mainly left-sided localized aneurysms, regions of wall thinning with segmental akinesis, in addition to typical electric and histological features known for arrhythmogenic right ventricular cardiomyopathy. We identified a homozygous truncation mutation, c.1660C>T (p.Q554X) in desmocollin-2 (DSC2), in affected individuals and determined a carrier frequency of this mutation of 9.4% (1 in 10.6) among 1535 Schmiedeleut Hutterites, suggesting a common founder in that subgroup. Immunohistochemistry of endomyocardial biopsy samples revealed altered expression of the truncated DSC2 protein at the intercalated discs but only minor changes in immunoreactivity of other desmosomal proteins. Recombinant expressed mutant DSC2 protein in cells confirmed a stable, partially processed truncated protein with cytoplasmic and membrane localization. CONCLUSIONS: A homozygous truncation mutation in DSC2 leads to a cardiac-restricted phenotype of an early onset biventricular arrhythmogenic cardiomyopathy. The truncated protein remains partially stable and localized at the intercalated discs. These data suggest that the processed DSC2 protein plays a role in maintaining desmosome integrity and function.