K. Õunap and K. Reinson discuss the biochemical and functional characterization of the NDUFC2 pathogenic variants identified in children with Leigh syndrome by R. Taylor and colleagues, in this issue ...of EMBO Molecular Medicine
Mitochondrial complex I deficiency is associated with a wide range of clinical presentations, including Leigh syndrome. Its genetic causes are heterogeneous, with poor genotype–phenotype correlation. It is impossible to identify the genetic defect of complex I deficiency using clinical observation and metabolic/imaging studies alone. As a result, whole‐exome sequencing (WES) is increasingly used in clinical work to identify an underlying genetic defect causing the disease. The article in this issue of EMBO Molecular Medicine by Alahmad et al (2020) is timely and valuable, as it expands on the genotype of mitochondrial complex I deficiency by identifying and characterising pathogenic variants of the NDUFC2 gene in children with Leigh syndrome.
Silver-Russell syndrome (SRS) and Beckwith-Wiedemann syndrome (BWS) are 2 clinically opposite growth-affecting disorders belonging to the group of congenital imprinting disorders. The expression of ...both syndromes usually depends on the parental origin of the chromosome in which the imprinted genes reside. SRS is characterized by severe intrauterine and postnatal growth retardation with various additional clinical features such as hemihypertrophy, relative macrocephaly, fifth finger clinodactyly, and triangular facies. BWS is an overgrowth syndrome with many additional clinical features such as macroglossia, organomegaly, and an increased risk of childhood tumors. Both SRS and BWS are clinically and genetically heterogeneous, and for clinical diagnosis, different diagnostic scoring systems have been developed. Six diagnostic scoring systems for SRS and 4 for BWS have been previously published. However, neither syndrome has common consensus diagnostic criteria yet. Most cases of SRS and BWS are associated with opposite epigenetic or genetic abnormalities in the 11p15 chromosomal region leading to opposite imbalances in the expression of imprinted genes. SRS is also caused by maternal uniparental disomy 7, which is usually identified in 5-10% of the cases, and is therefore the first imprinting disorder that affects 2 different chromosomes. In this review, we describe in detail the clinical diagnostic criteria and scoring systems as well as molecular causes in both SRS and BWS.
The SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) protein syntaxin-5 (Stx5) is essential for Golgi transport. In humans, the STX5 mRNA encodes two protein isoforms, ...Stx5 Long (Stx5L) from the first starting methionine and Stx5 Short (Stx5S) from an alternative starting methionine at position 55. In this study, we identify a human disorder caused by a single missense substitution in the second starting methionine (p.M55V), resulting in complete loss of the short isoform. Patients suffer from an early fatal multisystem disease, including severe liver disease, skeletal abnormalities and abnormal glycosylation. Primary human dermal fibroblasts isolated from these patients show defective glycosylation, altered Golgi morphology as measured by electron microscopy, mislocalization of glycosyltransferases, and compromised ER-Golgi trafficking. Measurements of cognate binding SNAREs, based on biotin-synchronizable forms of Stx5 (the RUSH system) and Förster resonance energy transfer (FRET), revealed that the short isoform of Stx5 is essential for intra-Golgi transport. Alternative starting codons of Stx5 are thus linked to human disease, demonstrating that the site of translation initiation is an important new layer of regulating protein trafficking.
We studied galactose supplementation in SLC35A2-congenital disorder of glycosylation (SLC35A2-CDG), caused by monoallelic pathogenic variants in SLC35A2 (Xp11.23), encoding the endoplasmic reticulum ...(ER) and Golgi UDP-galactose transporter. Patients present with epileptic encephalopathy, developmental disability, growth deficiency, and dysmorphism.
Ten patients with SLC35A2-CDG were supplemented with oral D-galactose for 18 weeks in escalating doses up to 1.5 g/kg/day. Outcome was assessed using the Nijmegen Pediatric CDG Rating Scale (NPCRS, ten patients) and by glycomics (eight patients).
SLC35A2-CDG patients demonstrated improvements in overall Nijmegen Pediatric CDG Rating Scale (NPCRS) score (P = 0.008), the current clinical assessment (P = 0.007), and the system specific involvement (P = 0.042) domains. Improvements were primarily in growth and development with five patients resuming developmental progress, which included postural control, response to stimuli, and chewing and swallowing amelioration. Additionally, there were improvements in gastrointestinal symptoms and epilepsy. One patient in our study did not show any clinical improvement. Galactose supplementation improved patients' glycosylation with decreased ratios of incompletely formed to fully formed glycans (M-gal/disialo, P = 0.012 and monosialo/disialo, P = 0.017) and increased levels of a fully galactosylated N-glycan (P = 0.05).
Oral D-galactose supplementation results in clinical and biochemical improvement in SLC35A2-CDG. Galactose supplementation may partially overcome the Golgi UDP-galactose deficiency and improves galactosylation. Oral galactose is well tolerated and shows promise as dietary therapy.
Germline pathogenic variants in chromatin-modifying enzymes are a common cause of pediatric developmental disorders. These enzymes catalyze reactions that regulate epigenetic inheritance via histone ...post-translational modifications and DNA methylation. Cytosine methylation (5-methylcytosine 5mC) of DNA is the quintessential epigenetic mark, yet no human Mendelian disorder of DNA demethylation has yet been delineated. Here, we describe in detail a Mendelian disorder caused by the disruption of DNA demethylation. TET3 is a methylcytosine dioxygenase that initiates DNA demethylation during early zygote formation, embryogenesis, and neuronal differentiation and is intolerant to haploinsufficiency in mice and humans. We identify and characterize 11 cases of human TET3 deficiency in eight families with the common phenotypic features of intellectual disability and/or global developmental delay; hypotonia; autistic traits; movement disorders; growth abnormalities; and facial dysmorphism. Mono-allelic frameshift and nonsense variants in TET3 occur throughout the coding region. Mono-allelic and bi-allelic missense variants localize to conserved residues; all but one such variant occur within the catalytic domain, and most display hypomorphic function in an assay of catalytic activity. TET3 deficiency and other Mendelian disorders of the epigenetic machinery show substantial phenotypic overlap, including features of intellectual disability and abnormal growth, underscoring shared disease mechanisms.
Summary Background Muscle biopsy is the gold standard for diagnosis of mitochondrial disorders because of the lack of sensitive biomarkers in serum. Fibroblast growth factor 21 (FGF-21) is a growth ...factor with regulatory roles in lipid metabolism and the starvation response, and concentrations are raised in skeletal muscle and serum in mice with mitochondrial respiratory chain deficiencies. We investigated in a retrospective diagnostic study whether FGF-21 could be a biomarker for human mitochondrial disorders. Methods We assessed samples from adults and children with mitochondrial disorders or non-mitochondrial neurological disorders (disease controls) from seven study centres in Europe and the USA, and recruited healthy volunteers (healthy controls), matched for age where possible, from the same centres. We used ELISA to measure FGF-21 concentrations in serum or plasma samples (abnormal values were defined as >200 pg/mL). We compared these concentrations with values for lactate, pyruvate, lactate-to-pyruvate ratio, and creatine kinase in serum or plasma and calculated sensitivity, specificity, and positive and negative predictive values for all biomarkers. Findings We analysed serum or plasma from 67 patients (41 adults and 26 children) with mitochondrial disorders, 34 disease controls (22 adults and 12 children), and 74 healthy controls. Mean FGF-21 concentrations in serum were 820 (SD 1151) pg/mL in adult and 1983 (1550) pg/mL in child patients with respiratory chain deficiencies and 76 (58) pg/mL in healthy controls. FGF-21 concentrations were high in patients with mitochondrial disorders affecting skeletal muscle but not in disease controls, including those with dystrophies. In patients with abnormal FGF-21 concentrations in serum, the odds ratio of having a muscle-manifesting mitochondrial disease was 132·0 (95% CI 38·7–450·3). For the identification of muscle-manifesting mitochondrial disease, the sensitivity was 92·3% (95% CI 81·5–97·9%) and specificity was 91·7% (84·8–96·1%). The positive and negative predictive values for FGF-21 were 84·2% (95% CI 72·1–92·5%) and 96·1 (90·4–98·9%). The accuracy of FGF-21 to correctly identify muscle-manifesting respiratory chain disorders was better than that for all conventional biomarkers. The area under the receiver-operating-characteristic curve for FGF-21 was 0·95; by comparison, the values for other biomarkers were 0·83 lactate (p=0·037, 0·83 for pyruvate (p=0·015), 0·72 for the lactate-to-pyruvate ratio (p=0·0002), and 0·77 for creatine kinase (p=0·013). Interpretation Measurement of FGF-21 concentrations in serum identified primary muscle-manifesting respiratory chain deficiencies in adults and children and might be feasible as a first-line diagnostic test for these disorders to reduce the need for muscle biopsy. Funding Sigrid Jusélius Foundation, Jane and Aatos Erkko Foundation, Molecular Medicine Institute of Finland, University of Helsinki, Helsinki University Central Hospital, Academy of Finland, Novo Nordisk, Arvo and Lea Ylppö Foundation.
Clinical interpretation of genetic variants in the context of the patient's phenotype is becoming the largest component of cost and time expenditure for genome-based diagnosis of rare genetic ...diseases. Artificial intelligence (AI) holds promise to greatly simplify and speed genome interpretation by integrating predictive methods with the growing knowledge of genetic disease. Here we assess the diagnostic performance of Fabric GEM, a new, AI-based, clinical decision support tool for expediting genome interpretation.
We benchmarked GEM in a retrospective cohort of 119 probands, mostly NICU infants, diagnosed with rare genetic diseases, who received whole-genome or whole-exome sequencing (WGS, WES). We replicated our analyses in a separate cohort of 60 cases collected from five academic medical centers. For comparison, we also analyzed these cases with current state-of-the-art variant prioritization tools. Included in the comparisons were trio, duo, and singleton cases. Variants underpinning diagnoses spanned diverse modes of inheritance and types, including structural variants (SVs). Patient phenotypes were extracted from clinical notes by two means: manually and using an automated clinical natural language processing (CNLP) tool. Finally, 14 previously unsolved cases were reanalyzed.
GEM ranked over 90% of the causal genes among the top or second candidate and prioritized for review a median of 3 candidate genes per case, using either manually curated or CNLP-derived phenotype descriptions. Ranking of trios and duos was unchanged when analyzed as singletons. In 17 of 20 cases with diagnostic SVs, GEM identified the causal SVs as the top candidate and in 19/20 within the top five, irrespective of whether SV calls were provided or inferred ab initio by GEM using its own internal SV detection algorithm. GEM showed similar performance in absence of parental genotypes. Analysis of 14 previously unsolved cases resulted in a novel finding for one case, candidates ultimately not advanced upon manual review for 3 cases, and no new findings for 10 cases.
GEM enabled diagnostic interpretation inclusive of all variant types through automated nomination of a very short list of candidate genes and disorders for final review and reporting. In combination with deep phenotyping by CNLP, GEM enables substantial automation of genetic disease diagnosis, potentially decreasing cost and expediting case review.
Although protein hydroxylation is a relatively poorly characterized posttranslational modification, it has received significant recent attention following seminal work uncovering its role in oxygen ...sensing and hypoxia biology. Although the fundamental importance of protein hydroxylases in biology is becoming clear, the biochemical targets and cellular functions often remain enigmatic. JMJD5 is a "JmjC-only" protein hydroxylase that is essential for murine embryonic development and viability. However, no germline variants in JmjC-only hydroxylases, including JMJD5, have yet been described that are associated with any human pathology. Here we demonstrate that biallelic germline JMJD5 pathogenic variants are deleterious to JMJD5 mRNA splicing, protein stability, and hydroxylase activity, resulting in a human developmental disorder characterized by severe failure to thrive, intellectual disability, and facial dysmorphism. We show that the underlying cellular phenotype is associated with increased DNA replication stress and that this is critically dependent on the protein hydroxylase activity of JMJD5. This work contributes to our growing understanding of the role and importance of protein hydroxylases in human development and disease.
Despite a few recent reports of patients harboring truncating variants in NSD2, a gene considered critical for the Wolf–Hirschhorn syndrome (WHS) phenotype, the clinical spectrum associated with NSD2 ...pathogenic variants remains poorly understood.
We collected a comprehensive series of 18 unpublished patients carrying heterozygous missense, elongating, or truncating NSD2 variants; compared their clinical data to the typical WHS phenotype after pooling them with ten previously described patients; and assessed the underlying molecular mechanism by structural modeling and measuring methylation activity in vitro.
The core NSD2-associated phenotype includes mostly mild developmental delay, prenatal-onset growth retardation, low body mass index, and characteristic facial features distinct from WHS. Patients carrying missense variants were significantly taller and had more frequent behavioral/psychological issues compared with those harboring truncating variants. Structural in silico modeling suggested interference with NSD2’s folding and function for all missense variants in known structures. In vitro testing showed reduced methylation activity and failure to reconstitute H3K36me2 in NSD2 knockout cells for most missense variants.
NSD2 loss-of-function variants lead to a distinct, rather mild phenotype partially overlapping with WHS. To avoid confusion for patients, NSD2 deficiency may be named Rauch–Steindl syndrome after the delineators of this phenotype.
Results of clinical genomic testing must be reported in a clear, concise format to ensure they are understandable and interpretable. It is important laboratories are aware of the information which is ...essential to make sure the results are not open to misinterpretation. As genomic testing has continued to evolve over the past decade, the European Society of Human Genetics (ESHG) recommendations for reporting results of diagnostic genetic testing (biochemical, cytogenetic and molecular genetic) published in 2014 have been reviewed and updated to provide the genomic community with guidance on reporting unambiguous results.
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