Heimler Syndrome Is Caused by Hypomorphic Mutations in the Peroxisome-Biogenesis Genes PEX1 and PEX6.

Heimler syndrome (HS) is a rare recessive disorder characterized by sensorineural hearing loss (SNHL), amelogenesis imperfecta, nail abnormalities, and occasional or late-onset retinal pigmentation. We ascertained eight families affected by HS and, by using a whole-exome sequencing approach, identified biallelic mutations in PEX1 or PEX6 in six of them. Loss-of-function mutations in both genes are known causes of a spectrum of autosomal-recessive peroxisome-biogenesis disorders (PBDs), including Zellweger syndrome. PBDs are characterized by leukodystrophy, hypotonia, SNHL, retinopathy, and skeletal, craniofacial, and liver abnormalities. We demonstrate that each HS-affected family has at least one hypomorphic allele that results in extremely mild peroxisomal dysfunction. Although individuals with HS share some subtle clinical features found in PBDs, the diagnosis was not suggested by routine blood and skin fibroblast analyses used to detect PBDs. In conclusion, our findings define HS as a mild PBD, expanding the pleiotropy of mutations in PEX1 and PEX6.

Mutational Spectrum in Holoprosencephaly Shows That FGF is a New Major Signaling Pathway.

Holoprosencephaly (HPE) is the most common congenital cerebral malformation in humans, characterized by impaired forebrain cleavage and midline facial anomalies. It presents a high heterogeneity, both in clinics and genetics. We have developed a novel targeted next-generation sequencing (NGS) assay and screened a cohort of 257 HPE patients. Mutations with high confidence in their deleterious effect were identified in approximately 24% of the cases and were held for diagnosis, whereas variants of uncertain significance were identified in 10% of cases. This study provides a new classification of genes that are involved in HPE. SHH, ZIC2, and SIX3 remain the top genes in term of frequency with GLI2, and are followed by FGF8 and FGFR1. The three minor HPE genes identified by our study are DLL1, DISP1, and SUFU. Here, we demonstrate that fibroblast growth factor signaling must now be considered a major pathway involved in HPE. Interestingly, several cases of double mutations were found and argue for a polygenic inheritance of HPE. Altogether, it supports that the implementation of NGS in HPE diagnosis is required to improve genetic counseling.

Whole-exome-sequencing identifies mutations in histone acetyltransferase gene KAT6B in individuals with the Say-Barber-Biesecker variant of Ohdo syndrome.

Say-Barber-Biesecker-Young-Simpson syndrome (SBBYSS or Ohdo syndrome) is a multiple anomaly syndrome characterized by severe intellectual disability, blepharophimosis, and a mask-like facial appearance. A number of individuals with SBBYSS also have thyroid abnormalities and cleft palate. The condition usually occurs sporadically and is therefore presumed to be due in most cases to new dominant mutations. In individuals with SBBYSS, a whole-exome sequencing approach was used to demonstrate de novo protein-truncating mutations in the highly conserved histone acetyltransferase gene KAT6B (MYST4/MORF)) in three out of four individuals sequenced. Sanger sequencing was used to confirm truncating mutations of KAT6B, clustering in the final exon of the gene in all four individuals and in a further nine persons with typical SBBYSS. Where parental samples were available, the mutations were shown to have occurred de novo. During mammalian development KAT6B is upregulated specifically in the developing central nervous system, facial structures, and limb buds. The phenotypic features seen in the Qkf mouse, a hypomorphic Kat6b mutant, include small eyes, ventrally placed ears and long first digits that mirror the human phenotype. This is a further example of how perturbation of a protein involved in chromatin modification might give rise to a multisystem developmental disorder.

Recessive mutations in MCM4/PRKDC cause a novel syndrome involving a primary immunodeficiency and a disorder of DNA repair.

BACKGROUND: A study is presented of 10 children with a novel syndrome born to consanguineous parents from the Irish Traveller population. The syndrome is characterised by a natural killer (NK) cell deficiency, evidence of an atypical Fanconi's type DNA breakage disorder, and features of familial glucocorticoid deficiency (FGD). The NK cell deficiency probably accounts for the patients' recurrent viral illnesses. Molecular tests support a diagnosis of mosaic Fanconi's anaemia, but the patients do not present with any of the expected clinical features of the disorder. The symptomatic presentation of FGD was delayed in onset and may be a secondary phenotype. As all three phenotypes segregate together, the authors postulated that the NK cell deficiency, DNA repair disorder and FGD were caused by a single recessive genetic event. METHODS: Single-nucleotide polymorphism homozygosity mapping and targeted next-generation sequencing of 10 patients and 16 unaffected relatives. RESULTS: A locus for the syndrome was identified at 8p11.21-q11.22. Targeted resequencing of the candidate region revealed a homozygous mutation in MCM4/PRKDC in all 10 affected individuals. Consistent with the observed DNA breakage disorder, MCM4 and PRKDC are both involved in the ATM/ATR (ataxia-telangiectasia-mutated/ATM-Rad 3-related) DNA repair pathway, which is defective in patients with Fanconi's anaemia. Deficiency of PRKDC in mice has been shown to result in an abnormal NK cell physiology similar to that observed in these patients. CONCLUSION: Mutations in MCM4/PRKDC represent a novel cause of DNA breakage and NK cell deficiency. These findings suggest that clinicians should consider this disorder in patients with failure to thrive who develop pigmentation or who have recurrent infections.

Mutations in SOX2 cause anophthalmia.

A submicroscopic deletion containing SOX2 was identified at the 3q breakpoint in a child with t(3;11)(q26.3;p11.2) associated with bilateral anophthalmia. Subsequent SOX2 mutation analysis identified de novo truncating mutations of SOX2 in 4 of 35 (11%) individuals with anophthalmia. Both eyes were affected in all cases with an identified mutation.

ARF1 prevents aberrant type I interferon induction by regulating STING activation and recycling.

Type I interferon (IFN) signalling is tightly controlled. Upon recognition of DNA by cyclic GMP-AMP synthase (cGAS), stimulator of interferon genes (STING) translocates along the endoplasmic reticulum (ER)-Golgi axis to induce IFN signalling. Termination is achieved through autophagic degradation or recycling of STING by retrograde Golgi-to-ER transport. Here, we identify the GTPase ADP-ribosylation factor 1 (ARF1) as a crucial negative regulator of cGAS-STING signalling. Heterozygous ARF1 missense mutations cause a previously unrecognized type I interferonopathy associated with enhanced IFN-stimulated gene expression. Disease-associated, GTPase-defective ARF1 increases cGAS-STING dependent type I IFN signalling in cell lines and primary patient cells. Mechanistically, mutated ARF1 perturbs mitochondrial morphology, causing cGAS activation by aberrant mitochondrial DNA release, and leads to accumulation of active STING at the Golgi/ERGIC due to defective retrograde transport. Our data show an unexpected dual role of ARF1 in maintaining cGAS-STING homeostasis, through promotion of mitochondrial integrity and STING recycling.

Identification of a mutation in LARS as a novel cause of infantile hepatopathy.

Infantile hepatopathies are life-threatening liver disorders that manifest in the first few months of life. We report on a consanguineous Irish Traveller family that includes six individuals presenting with acute liver failure in the first few months of life. Additional symptoms include anaemia, renal tubulopathy, developmental delay, seizures, failure to thrive and deterioration of liver function with minor illness. The multisystem manifestations suggested a possible mitochondrial basis to the disorder. However, known causes of childhood liver failure and mitochondrial disease were excluded in this family by biochemical, metabolic and genetic analyses. We aimed to identify the underlying risk gene using homozygosity mapping and whole exome sequencing. SNP homozygosity mapping identified a candidate locus at 5q31.3-q33.1. Whole exome sequencing identified 1 novel homozygous missense mutation within the 5q31.3-q33.1 candidate region that segregated with the hepatopathy. The candidate mutation is located in the LARS gene which encodes a cytoplasmic leucyl-tRNA synthetase enzyme responsible for exclusively attaching leucine to its cognate tRNA during protein translation. Knock-down of LARS in HEK293 cells did not impact on mitochondrial function even when the cells were put under physiological stress. The molecular studies confirm the findings of the patients' biochemical and genetic analyses which show that the hepatopathy is not a mitochondrial-based dysfunction problem, despite clinical appearances. This study highlights the clinical utility of homozygosity mapping and exome sequencing in diagnosing recessive liver disorders. It reports mutation of a cytoplasmic aminoacyl-tRNA synthetase enzyme as a possible novel cause of infantile hepatopathy and underscores the need to consider mutations in LARS in patients with liver disease and multisystem presentations.

First implication of STRA6 mutations in isolated anophthalmia, microphthalmia, and coloboma: a new dimension to the STRA6 phenotype.

Microphthalmia, anophthalmia, and coloboma (MAC) are structural congenital eye malformations that cause a significant proportion of childhood visual impairments. Several disease genes have been identified but do not account for all MAC cases, suggesting that additional risk loci exist. We used single nucleotide polymorphism (SNP) homozygosity mapping (HM) and targeted next-generation sequencing to identify the causative mutation for autosomal recessive isolated colobomatous microanophthalmia (MCOPCB) in a consanguineous Irish Traveller family. We identified a double-nucleotide polymorphism (g.1157G>A and g.1156G>A; p.G304K) in STRA6 that was homozygous in all of the MCOPCB patients. The STRA6 p.G304K mutation was subsequently detected in additional MCOPCB patients, including one individual with Matthew-Wood syndrome (MWS; MCOPS9). STRA6 encodes a transmembrane receptor involved in vitamin A uptake, a process essential to eye development and growth. We have shown that the G304K mutant STRA6 protein is mislocalized and has severely reduced vitamin A uptake activity. Furthermore, we reproduced the MCOPCB phenotype in a zebrafish disease model by inhibiting retinoic acid (RA) synthesis, suggesting that diminished RA levels account for the eye malformations in STRA6 p.G304K patients. The current study demonstrates that STRA6 mutations can cause isolated eye malformations in addition to the congenital anomalies observed in MWS.

A novel locus for episodic ataxia:UBR4 the likely candidate.

Episodic ataxias (EAs) are rare neurological channelopathies that are characterized by spells of imbalance and a lack of co-ordination. There are seven clinically recognized EAs and multiple isolated cases. Five disease-causing genes have been identified to date. We describe a novel form of autosomal dominant EA in a large three-generation Irish family. This form of EA presents in early childhood with periods of unsteadiness generalized weakness and slurred speech during an attack, which may be triggered by physical tiredness or stress. Linkage analysis undertaken in 13 related individuals identified a single disease locus (1p36.13-p34.3) with a LOD score of 3.29. Exome sequencing was performed. Following data analysis, which included presence/absence within the linkage peak, two candidate variants were identified. These are located in the HSPG2 and UBR4 genes. UBR4 is an ubiquitin ligase protein that is known to interact with calmodulin, a Ca(2+) protein, in the cytoplasm. It also co-localizes with ITPR1 a calcium release channel that is a major determinant of mammal co-ordination. Although UBR4 is not an ion channel gene, the potential for disrupted Ca(2+) control within neuronal cells highlights its potential for a role in this form of EA.

Thymidylate synthase repeat polymorphisms and risk of neural tube defects in a population from the northern United Kingdom.

BACKGROUND: A 28-bp repeat polymorphism in the 5'UTR of the thymidylate synthase (TYMS) gene represents a candidate risk factor for neural tube defects (NTDs) due to involvement in folate-dependent homocysteine metabolism. Non-Hispanic, white, U.S. citizens carrying at least one 2x 28-bp repeat allele have recently been shown to be at a four-fold increased risk of spina bifida (SB). We investigated the association between this polymorphism and risk of NTD in families affected by NTDs and controls from the northern United Kingdom (UK). METHODS: PCR was performed on genomic DNA extracted from blood or mouth swabs of family members affected by NTDs (mothers, fathers, and cases), and unaffected controls (mothers and infants) to determine the number of 28-bp repeat units within the promoter region of TYMS. Case-control and TDT analyses of the influence of TYMS genotype on risk of NTD, or NTD pregnancy, were conducted. RESULTS: Odds ratio (OR) analysis indicated that individuals carrying the 2x 28-bp repeat allele either in homozygous or heterozygous form, are not at increased risk of NTDs, or of having an NTD affected pregnancy. Control population allele frequencies are seen to be markedly different between the U.S. controls and those in this study. CONCLUSIONS: TYMS polymorphism appears to be not universally associated with NTD risk across Caucasian samples. The elevated risk of spina bifida in U.S. samples appears to be driven by an unusually low risk allele (2x 28 bp) frequency in control samples. Family based (TDT) testing of U.S. samples is therefore advocated.