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Neonatal diabetes is caused by single gene mutations reducing pancreatic [beta] cell number or impairing [beta] cell function. Understanding the genetic basis of rare diabetes subtypes highlights fundamental biological processes in [beta] cells. We identified 6 patients from 5 families with homozygous mutations in the YIPF5 gene, which is involved in trafficking between the endoplasmic reticulum (ER) and the Golgi. All patients had neonatal/early-onset diabetes, severe microcephaly, and epilepsy. YIPF5 is expressed during human brain development, in adult brain and pancreatic islets. We used 3 human [beta] cell models (YIPF5 silencing in EndoC-[beta]H1 cells, YIPF5 knockout and mutation knockin in embryonic stem cells, and patient-derived induced pluripotent stem cells) to investigate the mechanism through which YIPF5 loss of function affects [beta] cells. Loss of YIPF5 function in stem cell-derived islet cells resulted in proinsulin retention in the ER, marked ER stress, and p cell failure. Partial YIPF5 silencing in EndoC-[beta]H1 cells and a patient mutation in stem cells increased the [beta] cell sensitivity to ER stress-induced apoptosis. We report recessive YIPF5 mutations as the genetic cause of a congenital syndrome of microcephaly, epilepsy, and neonatal/early-onset diabetes, highlighting a critical role of YIPF5 in [beta] cells and neurons. We believe this is the first report of mutations disrupting the ER-to-Golgi trafficking, resulting in diabetes., Introduction Neonatal diabetes mellitus develops before 6 months of age and is caused by reduced pancreatic [beta] cell number (reduced formation/increased destruction) or impaired [beta] cell function. Previous studies have [...]

In silico predictive tools can help determine the pathogenicity of variants. The 2015 American College of Medical Genetics and Genomics (ACMG) guidelines recommended that scores from these tools can be used as supporting evidence of pathogenicity. A subsequent publication by the ClinGen Sequence Variant Interpretation Working Group suggested that high scores from some tools were sufficiently predictive to be used as moderate or strong evidence of pathogenicity. REVEL is a widely used metapredictor that uses the scores of 13 individual in silico tools to calculate the pathogenicity of missense variants. Its ability to predict missense pathogenicity has been assessed extensively; however, no study has previously tested whether its performance is affected by whether the missense variant acts via a loss-of-function (LoF) or gain-of-function (GoF) mechanism. We used a highly curated dataset of 66 confirmed LoF and 65 confirmed GoF variants to evaluate whether this affected the performance of REVEL. 98% of LoF and 100% of GoF variants met the author-recommended REVEL threshold of 0.5 for pathogenicity, while 89% of LoF and 88% of GoF variants exceeded the 0.75 threshold. However, while 55% of LoF variants met the threshold recommended for a REVEL score to count as strong evidence of pathogenicity from the ACMG guidelines (0.932), only 35% of GoF variants met this threshold (P = 0.0352). GoF variants are therefore less likely to receive the highest REVEL scores which would enable the REVEL score to be used as strong evidence of pathogenicity. This has implications for classification with the ACMG guidelines as GoF variants are less likely to meet the criteria for pathogenicity. [ABSTRACT FROM AUTHOR]

ONECUT1 (also known as HNF6) is a transcription factor involved in pancreatic development and β-cell function. Recently, biallelic variants in ONECUT1 were reported as a cause of neonatal diabetes mellitus (NDM) in two subjects, and missense monoallelic variants were associated with type 2 diabetes and possibly maturity-onset diabetes of the young (MODY). Here we examine the role of ONECUT1 variants in NDM, MODY, and type 2 diabetes in large international cohorts of subjects with monogenic diabetes and >400,000 subjects from UK Biobank. We identified a biallelic frameshift ONECUT1 variant as the cause of NDM in one individual. However, we found no enrichment of missense or null ONECUT1 variants among 484 individuals clinically suspected of MODY, in whom all known genes had been excluded. Finally, using a rare variant burden test in the UK Biobank European cohort, we identified a significant association between heterozygous ONECUT1 null variants and type 2 diabetes (P = 0.006) but did not find an association between missense variants and type 2 diabetes. Our results confirm biallelic ONECUT1 variants as a cause of NDM and highlight monoallelic null variants as a risk factor for type 2 diabetes. These findings confirm the critical role of ONECUT1 in human β-cell function. Article Highlights: We confirmed homozygous ONECUT1 variants as causative for neonatal diabetes with the identification of a third case. Rare heterozygous ONECUT1 variants were not enriched in a cohort of 484 individuals clinically suspected of having maturity-onset diabetes of the young. Heterozygous null ONECUT1 variants are significantly associated with type 2 diabetes in the UK Biobank European population. No association was observed between heterozygous ONECUT1 missense variants and type 2 diabetes in UK Biobank. [ABSTRACT FROM AUTHOR]

Maturity-onset diabetes of the young (MODY) is an autosomal dominant form of monogenic diabetes, reported to be caused by variants in 16 genes. Concern has been raised about whether variants in BLK (MODY11), KLF11 (MODY7), and PAX4 (MODY9) cause MODY. We examined variant-level genetic evidence (cosegregation with diabetes and frequency in population) for published putative pathogenic variants in these genes and used burden testing to test gene-level evidence in a MODY cohort (n = 1,227) compared with a control population (UK Biobank [n = 185,898]). For comparison we analyzed well-established causes of MODY, HNF1A, and HNF4A. The published variants in BLK, KLF11, and PAX4 showed poor cosegregation with diabetes (combined logarithm of the odds [LOD] scores ≤1.2), compared with HNF1A and HNF4A (LOD scores >9), and are all too common to cause MODY (minor allele frequency >4.95 × 10-5). Ultra-rare missense and protein-truncating variants (PTV) were not enriched in a MODY cohort compared with the UK Biobank population (PTV P > 0.05, missense P > 0.1 for all three genes) while HNF1A and HNF4A were enriched (P < 10-6). Findings of sensitivity analyses with different population cohorts supported our results. Variant and gene-level genetic evidence does not support BLK, KLF11, or PAX4 as a cause of MODY. They should not be included in MODY diagnostic genetic testing. [ABSTRACT FROM AUTHOR]

Identifying copy number variants (CNVs) can provide diagnoses to patients and provide important biological insights into human health and disease. Current exome and targeted sequencing approaches cannot detect clinically and biologically-relevant CNVs outside their target area. We present SavvyCNV, a tool which uses off-target read data from exome and targeted sequencing data to call germline CNVs genome-wide. Up to 70% of sequencing reads from exome and targeted sequencing fall outside the targeted regions. We have developed a new tool, SavvyCNV, to exploit this 'free data' to call CNVs across the genome. We benchmarked SavvyCNV against five state-of-the-art CNV callers using truth sets generated from genome sequencing data and Multiplex Ligation-dependent Probe Amplification assays. SavvyCNV called CNVs with high precision and recall, outperforming the five other tools at calling CNVs genome-wide, using off-target or on-target reads from targeted panel and exome sequencing. We then applied SavvyCNV to clinical samples sequenced using a targeted panel and were able to call previously undetected clinically-relevant CNVs, highlighting the utility of this tool within the diagnostic setting. SavvyCNV outperforms existing tools for calling CNVs from off-target reads. It can call CNVs genome-wide from targeted panel and exome data, increasing the utility and diagnostic yield of these tests. SavvyCNV is freely available at https://github.com/rdemolgen/SavvySuite. Author summary: We have created SavvyCNV, a new tool for calling genetic variants. Large regions of the genome can be deleted or duplicated–these variants can have important consequences, for example causing a patient's genetic disease. However, many standard genetic tests only target a small fraction of the genome and will miss variants outside of these regions. Therefore, we developed a tool to exploit sequencing data which falls outside of these regions (due to flaws in the targeting process) to call large deletions and duplications. This allows large deletions and duplications to be detected anywhere in the genome. Researchers and diagnostic laboratories can use this tool to discover more genetic variants by re-analysing their sequencing data. [ABSTRACT FROM AUTHOR]

Aims Methods Results Conclusions Neonatal diabetes is a monogenic condition which can be the presenting feature of complex syndromes. The aim of this study was to identify novel genetic causes of neonatal diabetes with neurological features including developmental delay and epilepsy.We performed genome sequencing in 27 individuals with neonatal diabetes plus epilepsy and/or developmental delay of unknown genetic cause. Replication studies were performed in 123 individuals with diabetes diagnosed aged ≤1 year without a known genetic cause using targeted next‐generation sequencing.Three individuals, all diagnosed with diabetes in the first week of life, shared a rare homozygous missense variant, p.(Arg327Gln), in TARS2. Replication studies identified the same homozygous variant in a fourth individual diagnosed with diabetes at 1 year. One proband had epilepsy, one had development delay and two had both.Biallelic TARS2 variants cause a mitochondrial encephalopathy (COXPD‐21) characterised by severe hypotonia, epilepsy and developmental delay. Diabetes is not a known feature of COXPD‐21. Current evidence suggests that the p.(Arg327Gln) variant disrupts TARS2's regulation of the mTORC1 pathway which is essential for β‐cells.Our findings establish the homozygous p.(Arg327Gln) TARS2 variant as a novel cause of syndromic neonatal diabetes and uncover a role for TARS2 in pancreatic β‐cells. [ABSTRACT FROM AUTHOR]

Mesencephalic astrocyte-derived neurotrophic factor (MANF) is an endoplasmic reticulum (ER)-resident protein that plays a crucial role in attenuating ER stress responses. Although MANF is indispensable for the survival and function of mouse β-cells, its precise role in human β-cell development and function is unknown. In this study, we show that lack of MANF in humans results in diabetes due to increased ER stress, leading to impaired β-cell function. We identified two patients from different families with childhood diabetes and a neurodevelopmental disorder associated with homozygous loss-of-function mutations in the MANF gene. To study the role of MANF in human β-cell development and function, we knocked out the MANF gene in human embryonic stem cells and differentiated them into pancreatic endocrine cells. Loss of MANF induced mild ER stress and impaired insulin-processing capacity of β-cells in vitro. Upon implantation to immunocompromised mice, the MANF knockout grafts presented elevated ER stress and functional failure, particularly in recipients with diabetes. By describing a new form of monogenic neurodevelopmental diabetes syndrome caused by disturbed ER function, we highlight the importance of adequate ER stress regulation for proper human β-cell function and demonstrate the crucial role of MANF in this process. [ABSTRACT FROM AUTHOR]

Permanent neonatal diabetes mellitus (PNDM) is caused by reduced β-cell number or impaired β-cell function. Understanding of the genetic basis of this disorder highlights fundamental β-cell mechanisms. We performed trio genome sequencing for 44 patients with PNDM and their unaffected parents to identify causative de novo variants. Replication studies were performed in 188 patients diagnosed with diabetes before 2 years of age without a genetic diagnosis. EIF2B1 (encoding the eIF2B complex α subunit) was the only gene with novel de novo variants (all missense) in at least three patients. Replication studies identified two further patients with de novo EIF2B1 variants. In addition to having diabetes, four of five patients had hepatitis-like episodes in childhood. The EIF2B1 de novo mutations were found to map to the same protein surface. We propose that these variants render the eIF2B complex insensitive to eIF2 phosphorylation, which occurs under stress conditions and triggers expression of stress response genes. Failure of eIF2B to sense eIF2 phosphorylation likely leads to unregulated unfolded protein response and cell death. Our results establish de novo EIF2B1 mutations as a novel cause of permanent diabetes and liver dysfunction. These findings confirm the importance of cell stress regulation for β-cells and highlight EIF2B1's fundamental role within this pathway. [ABSTRACT FROM AUTHOR]

The syntheses of a series of 1‐phenyl‐5‐phosphino 1,2,3‐triazoles are disclosed, within which, the phosphorus atom (at the 5‐position of a triazole) is appended by one, two or three triazole motifs, and the valency of the phosphorus(III) atom is completed by two, one or zero ancillary (phenyl or cyclohexyl) groups respectively. This series of phosphines was compared with tricyclohexylphosphine and triphenylphosphine to study the effect of increasing the number of triazoles appended to the central phosphorus atom from zero to three triazoles. Gold(I) chloride complexes of the synthesised ligands were prepared and analysed by techniques including single‐crystal X‐ray diffraction structure determination. Gold(I) complexes were also prepared from 1‐(2,6‐dimethoxy)‐phenyl‐5‐dicyclohexyl‐phosphino 1,2,3‐triazole and 1‐(2,6‐dimethoxy)‐phenyl‐5‐diphenyl‐phosphino 1,2,3‐triazole ligands. The crystal structures thus obtained were examined using the SambVca (2.0) web tool and percentage buried volumes determined. The effectiveness of these gold(I) chloride complexes to serve as precatalysts for alkyne hydration were assessed. Furthermore, the regioselectivity of hydration of but‐1‐yne‐1,4‐diyldibenzene was probed. [ABSTRACT FROM AUTHOR]

Identifying new causes of permanent neonatal diabetes (PNDM) (diagnosis <6 months) provides important insights into β-cell biology. Patients with Down syndrome (DS) resulting from trisomy 21 are four times more likely to have childhood diabetes with an intermediate HLA association. It is not known whether DS can cause PNDM. We found that trisomy 21 was seven times more likely in our PNDM cohort than in the population (13 of 1,522 = 85 of 10,000 observed vs. 12.6 of 10,000 expected) and none of the 13 DS-PNDM patients had a mutation in the known PNDM genes that explained 82.9% of non-DS PNDM. Islet autoantibodies were present in 4 of 9 DS-PNDM patients, but DS-PNDM was not associated with polygenic susceptibility to type 1 diabetes (T1D). We conclude that trisomy 21 is a cause of autoimmune PNDM that is not HLA associated. We propose that autoimmune diabetes in DS is heterogeneous and includes coincidental T1D that is HLA associated and diabetes caused by trisomy 21 that is not HLA associated. [ABSTRACT FROM AUTHOR]

Physical stress, including high temperatures, may damage the central metabolic nicotinamide nucleotide cofactors [NAD(P)H], generating toxic derivatives [NAD(P)HX]. The highly conserved enzyme NAD(P)HX dehydratase (NAXD) is essential for intracellular repair of NAD(P)HX. Here we present a series of infants and children who suffered episodes of febrile illness-induced neurodegeneration or cardiac failure and early death. Whole-exome or whole-genome sequencing identified recessive NAXD variants in each case. Variants were predicted to be potentially deleterious through in silico analysis. Reverse-transcription PCR confirmed altered splicing in one case. Subject fibroblasts showed highly elevated concentrations of the damaged cofactors S-NADHX, R-NADHX and cyclic NADHX. NADHX accumulation was abrogated by lentiviral transduction of subject cells with wild-type NAXD. Subject fibroblasts and muscle biopsies showed impaired mitochondrial function, higher sensitivity to metabolic stress in media containing galactose and azide, but not glucose, and decreased mitochondrial reactive oxygen species production. Recombinant NAXD protein harbouring two missense variants leading to the amino acid changes p.(Gly63Ser) and p.(Arg608Cys) were thermolabile and showed a decrease in Vmax and increase in KM for the ATP-dependent NADHX dehydratase activity. This is the first study to identify pathogenic variants in NAXD and to link deficient NADHX repair with mitochondrial dysfunction. The results show that NAXD deficiency can be classified as a metabolite repair disorder in which accumulation of damaged metabolites likely triggers devastating effects in tissues such as the brain and the heart, eventually leading to early childhood death. [ABSTRACT FROM AUTHOR]

Summary Current tools to annotate the predicted effect of genetic variants are heavily biased towards protein-coding sequence. Variants outside of these regions may have a large impact on protein expression and/or structure and can lead to disease, but this effect can be challenging to predict. Consequently, these variants are poorly annotated using standard tools. We have developed a plugin to the Ensembl Variant Effect Predictor, the UTRannotator, that annotates variants in 5 ′ untranslated regions (5 ′ UTR) that create or disrupt upstream open reading frames. We investigate the utility of this tool using the ClinVar database, providing an annotation for 31.9% of all 5 ′ UTR (likely) pathogenic variants, and highlighting 31 variants of uncertain significance as candidates for further follow-up. We will continue to update the UTRannotator as we gain new knowledge on the impact of variants in UTRs. Availability and implementation UTRannotator is freely available on Github: https://github.com/ImperialCardioGenetics/UTRannotator. Supplementary information Supplementary data are available at Bioinformatics online. [ABSTRACT FROM AUTHOR]

Infancy-onset diabetes, microcephaly, PDIA6, polycystic kidney disease, whole genome sequencing, transcript Keywords: infancy-onset diabetes; microcephaly; PDIA6; polycystic kidney disease; transcript; whole genome sequencing EN infancy-onset diabetes microcephaly PDIA6 polycystic kidney disease transcript whole genome sequencing 457 458 2 10/13/22 20221101 NES 221101 We report a second patient with intrauterine growth retardation, congenital polycystic kidney disease, infancy-onset diabetes, microcephaly, and liver fibrosis caused by a homozygous PDIA6 loss-of-function variant. A biallelic loss-of-function PDIA6 variant in a second patient with polycystic kidney disease, infancy-onset diabetes, and microcephaly. [Extracted from the article]

Satellite infrared sounders are invaluable tools for making observations of the structure of the atmosphere. They provide much of the observational data used to initialize atmospheric models, especially in regions that do not have extensive surface-based observing systems, such as oceans. However, information is lacking in the presence of cloud, as the cloud layer is opaque to infrared radiation. This means that where information is most desired (such as in a developing storm) it is often in the shortest supply. In order to explore the mathematics of assimilating data from cloudy radiances, a study has been performed using an idealized single-column atmospheric model. The model simulates cloud development in an atmosphere with vertical motion, allowing the characteristics of a 2D-Var data assimilation system using a single simulated infrared satellite observation taken multiple times to be studied. The strongly nonlinear nature of cloud formation poses a challenge for variational methods. The adjoint method produces an accurate gradient for the cost function and minimization is achieved using preconditioned conjugate gradients. The conditioning is poor and varies strongly with the atmospheric variables and the cost function has multiple minima, but acceptable results are achieved. The assimilation system is provided with a prior forecast simulated by adding random correlated Gaussian error to the truth. Assimilating observations comparable to those available from current geostationary satellites allows vertical motion to be retrieved with an error of less than a centimetre per second in most conditions. Moreover, evaluating the second derivative of the cost function at the minimum provides an estimate of the uncertainty in the retrieval. This allows atmospheric states that do not provide sufficient information for retrieval of vertical motion to be detected (such as a cloudless atmosphere or a non-moving opaque cloud layer in the upper troposphere). Retrieval is most accurate with upwards motion. [ABSTRACT FROM AUTHOR]

Neonatal diabetes is caused by single gene mutations affecting fundamental β-cell function pathways. 20% of cases of neonatal diabetes remain genetically unexplained. Our aim was to explore the genetic basis of a syndrome characterized by neonatal diabetes, microcephaly and epilepsy, present in 2 unrelated patients. The Turkish and Indian patients were born to consanguineous parents. Using whole genome sequencing, we found that they had homozygous likely deleterious variants (missense, p.(Ala181Val) and in-frame deletion p.(Lys106del)) in YIPF5. We then performed replication studies in 112 patients with neonatal diabetes by targeted next generation sequencing. These identified 2 homozygous YIPF5 mutations (p.(Trp218Arg) and p.(Ile98Ser)) in 3 patients (2 siblings) with early-onset diabetes, epilepsy and microcephaly. In functional studies, YIPF5 was expressed in human islets and fetal brain cortex, as evaluated by qPCR and in situ hybridization. Because YIPF5 is involved in endoplasmic reticulum (ER)-to-Golgi trafficking, we examined the impact of YIPF5 loss-of-function on β-cell survival during ER stress. YIPF5 was silenced in the human β-cell line EndoC-βH1 and in human islets. YIPF5 silencing sensitized β cells to apoptosis induced by the ER stressors thapsigargin and brefeldin A. The ER stressors enhanced CHOP, BiP and spliced XBP1 expression in YIPF5-depleted cells, indicating increased ER stress signaling. Expression of the proapoptotic proteins PUMA and DP5 was also enhanced by YIPF5 silencing. CHOP and DP5 knockdown partially protected YIPF5-deficient cells from apoptosis, suggesting that they mediate apoptosis. In conclusion, homozygous loss-of-function mutations in YIPF5 are a novel cause of early-onset diabetes, microcephaly and epilepsy. This syndrome unveils a critical role of YIPF5 and ER-to-Golgi trafficking in the function and survival of human β-cells and neurons. Disclosure: M. Lytrivi: None. E. De Franco: Other Relationship; Self; Novo Nordisk Foundation. K.A. Patel: None. M. Igoillo Esteve: None. C. Cosentino: None. M.N. Wakeling: None. B. Haliloglu: None. M. Yildiz: None. T. Godbole: None. A.T. Hattersley: None. M. Cnop: None. Funding: Fonds Erasme for Medical Research; European Union (667191); Fonds National de la Recherche Scientifique; Brussels-Capital Region Innoviris [ABSTRACT FROM AUTHOR]

The article discusses the p.(Gly111Arg) ABCC8 variant as a founder mutation causing congenital hyperinsulinism in the Indian Agarwal community. The study identifies individuals with this variant, predominantly from the Agarwal community, and highlights the genetic homogeneity within this group due to clan exogamy and caste endogamy practices. The findings suggest that the p.(Gly111Arg) variant is a founder variant in the Agarwal community, enabling rapid prenatal screening and molecular diagnosis for congenital hyperinsulinism in this population. [Extracted from the article]

Neonatal diabetes is caused by single gene mutations reducing pancreatic β cell number or impairing β cell function. Understanding the genetic basis of rare diabetes subtypes highlights fundamental biological processes in β cells. We identified 6 patients from 5 families with homozygous mutations in the YIPF5 gene, which is involved in trafficking between the endoplasmic reticulum (ER) and the Golgi. All patients had neonatal/early-onset diabetes, severe microcephaly, and epilepsy. YIPF5 is expressed during human brain development, in adult brain and pancreatic islets. We used 3 human β cell models (YIPF5 silencing in EndoC-βH1 cells, YIPF5 knockout and mutation knockin in embryonic stem cells, and patient-derived induced pluripotent stem cells) to investigate the mechanism through which YIPF5 loss of function affects β cells. Loss of YIPF5 function in stem cell-derived islet cells resulted in proinsulin retention in the ER, marked ER stress, and β cell failure. Partial YIPF5 silencing in EndoC-βH1 cells and a patient mutation in stem cells increased the β cell sensitivity to ER stress-induced apoptosis. We report recessive YIPF5 mutations as the genetic cause of a congenital syndrome of microcephaly, epilepsy, and neonatal/early-onset diabetes, highlighting a critical role of YIPF5 in β cells and neurons. We believe this is the first report of mutations disrupting the ER-to-Golgi trafficking, resulting in diabetes. [ABSTRACT FROM AUTHOR]

Introduction: Biallelic PDX1 variants are a rare cause of isolated pancreatic agenesis and neonatal diabetes (NDM) without exocrine pancreatic insufficiency, with 17 cases reported in the literature., Research Design and Methods: To determine the phenotypic variability caused by this rare genetic aetiology, we investigated 19 individuals with NDM resulting from biallelic disease-causing PDX1 variants., Results: Of the 19 individuals, 8 (42%) were confirmed to have exocrine insufficiency requiring replacement therapy. Twelve individuals (63.2%) had extrapancreatic features, including 8 (42%) with conditions affecting the duodenum and/or hepatobiliary tract. Defects in duodenum development are consistent with previous Pdx1 ablation studies in mice which showed abnormal rostral duodenum development., Conclusions: Our findings show that recessive PDX1 variants can cause a syndromic form of NDM, highlighting the need for clinical assessment of extrapancreatic features in individuals with NDM caused by PDX1 variants., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2024. Re-use permitted under CC BY. Published by BMJ.)

Context: Hyperinsulinemic hypoglycemia (HI) can be the presenting feature of Kabuki syndrome (KS), which is caused by loss-of-function variants in KMT2D or KDM6A. As these genes play a critical role in maintaining methylation status in chromatin, individuals with pathogenic variants have a disease-specific epigenomic profile -an episignature., Objective: We evaluated the pathogenicity of three novel partial KDM6A duplications identified in three individuals presenting with neonatal-onset HI without typical features of KS at the time of genetic testing., Methods: Three different partial KDM6A duplications were identified by routine targeted next generation sequencing for HI and initially classified as variants of uncertain significance (VUS) as their location, and hence their impact on the gene, was not known. Whole genome sequencing (WGS) was undertaken to map the breakpoints of the duplications with DNA methylation profiling performed in two individuals to investigate the presence of a KS-specific episignature., Results: WGS confirmed the duplication in proband 1 as pathogenic as it caused a frameshift in the normal copy of the gene leading to a premature termination codon. The duplications identified in probands 2 and 3 did not alter the reading frame and therefore their significance remained uncertain after WGS. Subsequent DNA methylation profiling identified a KS-specific episignature in proband 2 but not in proband 3., Conclusions: Our findings confirm a role for KDM6A partial gene duplications in the etiology of KS and highlight the importance of performing in-depth molecular genetic analysis to properly assess the clinical significance of VUS's in the KDM6A gene., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Endocrine Society.)

We previously reported two siblings with inherited PD-1 deficiency who died from autoimmune pneumonitis at 3 and 11 years of age after developing other autoimmune manifestations, including type 1 diabetes (T1D). We report here two siblings, aged 10 and 11 years, with neonatal-onset T1D (diagnosed at the ages of 1 day and 7 wk), who are homozygous for a splice-site variant of CD274 (encoding PD-L1). This variant results in the exclusive expression of an alternative, loss-of-function PD-L1 protein isoform in overexpression experiments and in the patients' primary leukocytes. Surprisingly, cytometric immunophenotyping and single-cell RNA sequencing analysis on blood leukocytes showed largely normal development and transcriptional profiles across lymphoid and myeloid subsets in the PD-L1-deficient siblings, contrasting with the extensive dysregulation of both lymphoid and myeloid leukocyte compartments in PD-1 deficiency. Our findings suggest that PD-1 and PD-L1 are essential for preventing early-onset T1D but that, unlike PD-1 deficiency, PD-L1 deficiency does not lead to fatal autoimmunity with extensive leukocytic dysregulation., Competing Interests: Disclosures: C. Speake reported personal fees from Vertex Pharmaceuticals and GentiBio outside the submitted work. M.S. Anderson reported other from Merck, Inc. outside the submitted work. R.A. Oram reported grants from Randox, and personal fees from Sanofi, Provention Bio, and Janssen outside the submitted work; and that Randox are licensing knowhow relating to T1D polygenic scores from the University of Exeter. No other disclosures were reported., (© 2024 Johnson et al.)

A polysubstituted 3-aminoimidazo[5,1- b ]oxazol-6-ium framework has been accessed from a new nitrenoid reagent by a two-step ynamide annulation and imidazolium ring-formation sequence. Metalation with Au(I), Cu(I) and Ir(I) at the C2 position provides an L-shaped NHC ligand scaffold that has been validated in gold-catalysed alkyne hydration and arylative cyclisation reactions., (Copyright © 2024, Gillie et al.)

SLC4A10 is a plasma-membrane bound transporter that utilizes the Na+ gradient to drive cellular HCO3- uptake, thus mediating acid extrusion. In the mammalian brain, SLC4A10 is expressed in principal neurons and interneurons, as well as in epithelial cells of the choroid plexus, the organ regulating the production of CSF. Using next generation sequencing on samples from five unrelated families encompassing nine affected individuals, we show that biallelic SLC4A10 loss-of-function variants cause a clinically recognizable neurodevelopmental disorder in humans. The cardinal clinical features of the condition include hypotonia in infancy, delayed psychomotor development across all domains and intellectual impairment. Affected individuals commonly display traits associated with autistic spectrum disorder including anxiety, hyperactivity and stereotyped movements. In two cases isolated episodes of seizures were reported in the first few years of life, and a further affected child displayed bitemporal epileptogenic discharges on EEG without overt clinical seizures. While occipitofrontal circumference was reported to be normal at birth, progressive postnatal microcephaly evolved in 7 out of 10 affected individuals. Neuroradiological features included a relative preservation of brain volume compared to occipitofrontal circumference, characteristic narrow sometimes 'slit-like' lateral ventricles and corpus callosum abnormalities. Slc4a10 -/- mice, deficient for SLC4A10, also display small lateral brain ventricles and mild behavioural abnormalities including delayed habituation and alterations in the two-object novel object recognition task. Collapsed brain ventricles in both Slc4a10-/- mice and affected individuals suggest an important role of SLC4A10 in the production of the CSF. However, it is notable that despite diverse roles of the CSF in the developing and adult brain, the cortex of Slc4a10-/- mice appears grossly intact. Co-staining with synaptic markers revealed that in neurons, SLC4A10 localizes to inhibitory, but not excitatory, presynapses. These findings are supported by our functional studies, which show the release of the inhibitory neurotransmitter GABA is compromised in Slc4a10-/- mice, while the release of the excitatory neurotransmitter glutamate is preserved. Manipulation of intracellular pH partially rescues GABA release. Together our studies define a novel neurodevelopmental disorder associated with biallelic pathogenic variants in SLC4A10 and highlight the importance of further analyses of the consequences of SLC4A10 loss-of-function for brain development, synaptic transmission and network properties., (© The Author(s) 2023. Published by Oxford University Press on behalf of the Guarantors of Brain.)

Herein, we disclose a Ni-catalyzed 1,1-difunctionalization of unactivated terminal alkenes that enables the incorporation of two different heteroatom motifs across an olefin backbone, thus streamlining the access to α-aminoboronic acid derivatives from simple precursors. The method is characterized by its simplicity and generality across a wide number of coupling counterparts., Competing Interests: The authors declare no competing financial interest., (© 2023 American Chemical Society.)

Phosphatidylinositol 4-kinase IIIα (PI4KIIIα/PI4KA/OMIM:600286) is a lipid kinase generating phosphatidylinositol 4-phosphate (PI4P), a membrane phospholipid with critical roles in the physiology of multiple cell types. PI4KIIIα's role in PI4P generation requires its assembly into a heterotetrameric complex with EFR3, TTC7 and FAM126. Sequence alterations in two of these molecular partners, TTC7 (encoded by TTC7A or TCC7B) and FAM126, have been associated with a heterogeneous group of either neurological (FAM126A) or intestinal and immunological (TTC7A) conditions. Here we show that biallelic PI4KA sequence alterations in humans are associated with neurological disease, in particular hypomyelinating leukodystrophy. In addition, affected individuals may present with inflammatory bowel disease, multiple intestinal atresia and combined immunodeficiency. Our cellular, biochemical and structural modelling studies indicate that PI4KA-associated phenotypical outcomes probably stem from impairment of PI4KIIIα-TTC7-FAM126's organ-specific functions, due to defective catalytic activity or altered intra-complex functional interactions. Together, these data define PI4KA gene alteration as a cause of a variable phenotypical spectrum and provide fundamental new insight into the combinatorial biology of the PI4KIIIα-FAM126-TTC7-EFR3 molecular complex., (© The Author(s) (2021). Published by Oxford University Press on behalf of the Guarantors of Brain.)

Background: Large contiguous gene deletions at the distal end of the short arm of chromosome 9 result in the complex multi-organ condition chromosome 9p deletion syndrome.  A range of clinical features can result from these deletions with the most common being facial dysmorphisms and neurological impairment. Congenital hyperinsulinism is a rarely reported feature of the syndrome with the genetic mechanism for the dysregulated insulin secretion being unknown.  Methods: We studied the clinical and genetic characteristics of 12 individuals with chromosome 9p deletions who had a history of neonatal hypoglycaemia. Using off-target reads generated from targeted next-generation sequencing of the genes known to cause hyperinsulinaemic hypoglycaemia (n=9), or microarray analysis (n=3), we mapped the minimal shared deleted region on chromosome 9 in this cohort. Targeted sequencing was performed in three patients to search for a recessive mutation unmasked by the deletion. Results: In 10/12 patients with hypoglycaemia, hyperinsulinism was confirmed biochemically. A range of extra-pancreatic features were also reported in these patients consistent with the diagnosis of the Chromosome 9p deletion syndrome. The minimal deleted region was mapped to 7.2 Mb, encompassing 38 protein-coding genes. In silico analysis of these genes highlighted SMARCA2 and RFX3 as potential candidates for the hypoglycaemia. Targeted sequencing performed on three of the patients did not identify a second disease-causing variant within the minimal deleted region. Conclusions: This study identifies 9p deletions as an important cause of hyperinsulinaemic hypoglycaemia and increases the number of cases reported with 9p deletions and hypoglycaemia to 15 making this a more common feature of the syndrome than previously appreciated.  Whilst the precise genetic mechanism of the dysregulated insulin secretion could not be determined in these patients, mapping the deletion breakpoints highlighted potential candidate genes for hypoglycaemia within the deleted region., Competing Interests: No competing interests were disclosed., (Copyright: © 2020 Banerjee I et al.)

Permanent neonatal diabetes mellitus (PNDM) is caused by reduced β-cell number or impaired β-cell function. Understanding of the genetic basis of this disorder highlights fundamental β-cell mechanisms. We performed trio genome sequencing for 44 patients with PNDM and their unaffected parents to identify causative de novo variants. Replication studies were performed in 188 patients diagnosed with diabetes before 2 years of age without a genetic diagnosis. EIF2B1 (encoding the eIF2B complex α subunit) was the only gene with novel de novo variants (all missense) in at least three patients. Replication studies identified two further patients with de novo EIF2B1 variants. In addition to having diabetes, four of five patients had hepatitis-like episodes in childhood. The EIF2B1 de novo mutations were found to map to the same protein surface. We propose that these variants render the eIF2B complex insensitive to eIF2 phosphorylation, which occurs under stress conditions and triggers expression of stress response genes. Failure of eIF2B to sense eIF2 phosphorylation likely leads to unregulated unfolded protein response and cell death. Our results establish de novo EIF2B1 mutations as a novel cause of permanent diabetes and liver dysfunction. These findings confirm the importance of cell stress regulation for β-cells and highlight EIF2B1's fundamental role within this pathway., (© 2019 by the American Diabetes Association.)

Background: Large contiguous gene deletions at the distal end of the short arm of chromosome 9 result in the complex multi-organ condition chromosome 9p deletion syndrome.  A range of clinical features can result from these deletions with the most common being facial dysmorphisms and neurological impairment. Congenital hyperinsulinism is a rarely reported feature of the syndrome with the genetic mechanism for the dysregulated insulin secretion being unknown.  Methods: We studied the clinical and genetic characteristics of 12 individuals with chromosome 9p deletions who had a history of neonatal hypoglycaemia. Using off-target reads generated from targeted next-generation sequencing of the genes known to cause hyperinsulinaemic hypoglycaemia (n=9), or microarray analysis (n=3), we mapped the minimal shared deleted region on chromosome 9 in this cohort. Targeted sequencing was performed in three patients to search for a recessive mutation unmasked by the deletion. Results: In 10/12 patients with hypoglycaemia, hyperinsulinism was confirmed biochemically. A range of extra-pancreatic features were also reported in these patients consistent with the diagnosis of the Chromosome 9p deletion syndrome. The minimal deleted region was mapped to 7.2 Mb, encompassing 38 protein-coding genes. In silico analysis of these genes highlighted SMARCA2 and RFX3 as potential candidates for the hypoglycaemia. Targeted sequencing performed on three of the patients did not identify a second disease-causing variant within the minimal deleted region. Conclusions: This study identifies 9p deletions as an important cause of hyperinsulinaemic hypoglycaemia and increases the number of cases reported with 9p deletions and hypoglycaemia to 15 making this a more common feature of the syndrome than previously appreciated.  Whilst the precise genetic mechanism of the dysregulated insulin secretion could not be determined in these patients, mapping the deletion breakpoints highlighted potential candidate genes for hypoglycaemia within the deleted region., Competing Interests: No competing interests were disclosed., (Copyright: © 2019 Banerjee I et al.)

Multiple Nucleotide Variants (MNVs) are miscalled by the most widely utilised next generation sequencing analysis (NGS) pipelines, presenting the potential for missing diagnoses that would previously have been made by standard Sanger (dideoxy) sequencing. These variants, which should be treated as a single insertion-deletion mutation event, are commonly called as separate single nucleotide variants. This can result in misannotation, incorrect amino acid predictions and potentially false positive and false negative diagnostic results. This risk will be increased as confirmatory Sanger sequencing of Single Nucleotide variants (SNVs) ceases to be standard practice. Using simulated data and re-analysis of sequencing data from a diagnostic targeted gene panel, we demonstrate that the widely adopted pipeline, GATK best practices, results in miscalling of MNVs and that alternative tools can call these variants correctly. The adoption of calling methods that annotate MNVs correctly would present a solution for individual laboratories, however GATK best practices are the basis for important public resources such as the gnomAD database. We suggest integrating a solution into these guidelines would be the optimal approach., Competing Interests: No competing interests were disclosed., (Copyright: © 2019 Wakeling MN et al.)

Physical stress, including high temperatures, may damage the central metabolic nicotinamide nucleotide cofactors [NAD(P)H], generating toxic derivatives [NAD(P)HX]. The highly conserved enzyme NAD(P)HX dehydratase (NAXD) is essential for intracellular repair of NAD(P)HX. Here we present a series of infants and children who suffered episodes of febrile illness-induced neurodegeneration or cardiac failure and early death. Whole-exome or whole-genome sequencing identified recessive NAXD variants in each case. Variants were predicted to be potentially deleterious through in silico analysis. Reverse-transcription PCR confirmed altered splicing in one case. Subject fibroblasts showed highly elevated concentrations of the damaged cofactors S-NADHX, R-NADHX and cyclic NADHX. NADHX accumulation was abrogated by lentiviral transduction of subject cells with wild-type NAXD. Subject fibroblasts and muscle biopsies showed impaired mitochondrial function, higher sensitivity to metabolic stress in media containing galactose and azide, but not glucose, and decreased mitochondrial reactive oxygen species production. Recombinant NAXD protein harbouring two missense variants leading to the amino acid changes p.(Gly63Ser) and p.(Arg608Cys) were thermolabile and showed a decrease in Vmax and increase in KM for the ATP-dependent NADHX dehydratase activity. This is the first study to identify pathogenic variants in NAXD and to link deficient NADHX repair with mitochondrial dysfunction. The results show that NAXD deficiency can be classified as a metabolite repair disorder in which accumulation of damaged metabolites likely triggers devastating effects in tissues such as the brain and the heart, eventually leading to early childhood death.