Novel UBE3B mutations: report of eight patients with Kaufman oculocerebrofacial syndrome with additional clinical findings from a highly consanguineous population.

Biallelic mutations in UBE3B cause Kaufman oculocerebrofacial syndrome (KOS; OMIM 244450) with a wide range of clinical manifestations. In this study, we employed genetic analyses including homozygosity mapping, candidate gene sequencing, whole exome sequencing, and confirmatory Sanger sequencing on eight patients from three unrelated consanguineous families. Our analysis yielded three different novel variants in UBE3B : a missense substitution [NM_130466.4: c.2975C>T; (p.Pro992Leu)] in the HECT domain in family 1, a 3-bp deletion within exon 14 [c.1692_1694delCTC; (p.Ser565del)] leading to removal of a serine residue in family 2, and a splice donor site variant in intron eight of UBE3B (c.630 + 1G>T) in family 3. Blepharophimosis, telecanthus, ptosis, intellectual disability and abnormal lipid profile were similar to those found in previously reported KOS patients. Longitudinal follow-up revealed rather marfanoid body habitus of the patients in family 1. This study reports eight patients from Saudi Arabia with novel deleterious variants in UBE3B and adds to the phenotypic spectrum of KOS.

Towards a Cure for HARS Disease.

Histidyl-tRNA synthetase (HARS) ligates histidine to its cognate transfer RNA (tRNAHis). Mutations in HARS cause the human genetic disorders Usher syndrome type 3B (USH3B) and Charcot-Marie-Tooth syndrome type 2W (CMT2W). Treatment for these diseases remains symptomatic, and no disease specific treatments are currently available. Mutations in HARS can lead to destabilization of the enzyme, reduced aminoacylation, and decreased histidine incorporation into the proteome. Other mutations lead to a toxic gain-of-function and mistranslation of non-cognate amino acids in response to histidine codons, which can be rescued by histidine supplementation in vitro. We discuss recent advances in characterizing HARS mutations and potential applications of amino acid and tRNA therapy for future gene and allele specific therapy.

Histidine supplementation can escalate or rescue HARS deficiency in a Charcot-Marie-Tooth disease model.

Aminoacyl-tRNA synthetases are essential enzymes responsible for charging amino acids onto cognate tRNAs during protein synthesis. In histidyl-tRNA synthetase (HARS), autosomal dominant mutations V133F, V155G, Y330C and S356N in the HARS catalytic domain cause Charcot-Marie-Tooth disease type 2 W (CMT2W), while tRNA-binding domain mutation Y454S causes recessive Usher syndrome type IIIB. In a yeast model, all human HARS variants complemented a genomic deletion of the yeast ortholog HTS1 at high expression levels. CMT2W associated mutations, but not Y454S, resulted in reduced growth. We show mistranslation of histidine to glutamine and threonine in V155G and S356N but not Y330C mutants in yeast. Mistranslating V155G and S356N mutants lead to accumulation of insoluble proteins, which was rescued by histidine. Mutants V133F and Y330C showed the most significant growth defect and decreased HARS abundance in cells. Here, histidine supplementation led to insoluble protein aggregation and further reduced viability, indicating histidine toxicity associated with these mutants. V133F proteins displayed reduced thermal stability in vitro, which was rescued by tRNA. Our data will inform future treatment options for HARS patients, where histidine supplementation may either have a toxic or compensating effect depending on the nature of the causative HARS variant.

The Role of 3' to 5' Reverse RNA Polymerization in tRNA Fidelity and Repair.

The tRNAHis guanylyltransferase (Thg1) superfamily includes enzymes that are found in all three domains of life that all share the common ability to catalyze the 3' to 5' synthesis of nucleic acids. This catalytic activity, which is the reverse of all other known DNA and RNA polymerases, makes this enzyme family a subject of biological and mechanistic interest. Previous biochemical, structural, and genetic investigations of multiple members of this family have revealed that Thg1 enzymes use the 3' to 5' chemistry for multiple reactions in biology. Here, we describe the current state of knowledge regarding the catalytic features and biological functions that have been so far associated with Thg1 and its homologs. Progress toward the exciting possibility of utilizing this unusual protein activity for applications in biotechnology is also discussed.

Association of IL-13 rs20541 and rs1295686 variants with symptomatic asthma in a Saudi Arabian population.

OBJECTIVE: Interleukin 13 (IL-13) plays a critical pro-inflammatory role in asthma. Several single nucleotide polymorphisms (SNPs) are associated with asthma susceptibility in specific populations; however, further replicative studies in other ethnic groups are mandatory. METHODS: The association between IL-13 SNPs rs762534, rs20541, rs1295686, and rs1800925 (risk alleles A, A, T, and A, respectively) and asthma predisposition in a Saudi Arabian cohort was examined via a case-control cross-sectional study. RESULTS: The frequencies of alleles between asthmatics and control populations were significantly different for rs20541 and rs1295686 SNPs (p < 0.001), whereas the frequencies of genotypes between asthmatics and controls were significantly different only for rs20541. The association of the risk (minor) alleles with asthma was examined using the dominant genetic model. Individuals with at least one copy of the risk alleles A (for rs20541) and T (for rs1295686) had significantly greater odds of being asthmatic (OR = 2.13, 95% CI = 1.39-3.26, p < 0.0001; OR = 1.69, 95% CI = 1.12-2.54, p = 0.008) relative to their most common homozygous genotypes. On the other hand, the minor A alleles for rs762534 and rs1800925 were not significantly associated with asthma risk. Regarding haplotype association analysis, individuals with at least one copy of the minor "risk" allele for both rs20541 and rs1295686 (CATG and CATA, respectively) had greater odds of being asthmatic relative to CGCG haplotype; however, this trend was not statistically significant (p > 0.3). CONCLUSIONS: IL-13 minor T and A alleles for rs1295686 and rs20541, respectively, were associated with significantly higher risk of asthma in the Saudi Arabian population.

A novel anti-IL4Rα nanoparticle efficiently controls lung inflammation during asthma.

Drug resistance and the harmful side effects accompanying the prolonged corticosteroid treatment of chronic pulmonary diseases prompted the development of more specific anti-inflammatory approaches. Several strategies aiming to block IL4Rα, the receptor for a key pro-inflammatory pathway, were investigated. However, their efficiency was limited, mostly due to the systemic or subcutaneous route of administrations. In this paper, we examined the ability of an intranasal treatment with biocompatible nanoparticles targeting IL4Rα to control lung inflammation in ovalbumin (OVA)-sensitized mice. OVA-sensitized mice were treated with anti-IL4Rα-conjugated nanoparticles. The levels of pro-inflammatory cytokines in the lungs and broncho-alveolar lavage fluid (BALF) were determined using a cytokine array assay. The effects of nanoparticle treatment on the activation of lung inflammatory cells and their ability to proliferate and produce cytokines were determined using fluorescence-activated cell sorting (FACS) analysis. Lung inflammation was also monitored using immunohistochemical staining. Treatment with the anti-IL4Rα nanoparticles significantly decreased pro-inflammatory cytokine expression and release in BALF and airway lung tissue in mice. The numbers of lung tissue lymphocytes, neutrophils and eosinophils were also decreased. Interestingly, anti-IL4Rα nanoparticles deactivated CD4 and CD8 T cells in lung tissue and inhibited their ability to produce pro-inflammatory cytokines to a significantly lower level than the treatment with free anti-IL4Rα. Moreover, they induced a sustained low level of lung inflammation for 1 week following the last instillation compared with the treatment with free anti-IL4Rα antibodies. Together, this data suggested that the enhanced tissue penetrability and sustainability of these nanoparticles improved the strength and durability of the immunosuppressive effects of anti-IL4Rα.

KCNA4 deficiency leads to a syndrome of abnormal striatum, congenital cataract and intellectual disability.

BACKGROUND: Voltage-gated potassium channels are highly diverse proteins representing the most complex class of voltage-gated ion channels from structural and functional perspectives. Deficiency of these channels usually results in various human disorders. OBJECTIVES: To describe a novel autosomal recessive syndrome associated with KCNA4 deficiency leading to congenital cataract, abnormal striatum, intellectual disability and attention deficit hyperactivity disorder. METHODS: We used SNP arrays, linkage analyses, autozygosity mapping, whole-exome sequencing, RT-PCR and two-electrode voltage-clamp recording. RESULTS: We identified a missense variant (p.Arg89Gln) in KCNA4 in four patients from a consanguineous family manifesting a novel syndrome of congenital cataract, abnormal striatum, intellectual disability and attention deficit hyperactivity disorder. The variant was fully segregated with the disease and absent in 747 ethnically matched exomes. Xenopus oocytes were injected with human Kv1.4 wild-type mRNA, R89Q and WT/R89Q channels. The wild type had mean current amplitude that was significantly greater than those recorded from the cells expressing the same amount of mutant mRNA. Co-expression of the wild type and mutant mRNAs resulted in mean current amplitude that was significantly different from that of the wild type. RT-PCR indicated that KCNA4 is present in mouse brain, lens and retina. KCNA4 interacts with several molecules including synaptotagmin I, DLG1 and DLG2. The channel co-localises with cholinergic amacrine and rod bipolar cells in rats and is widely distributed in the central nervous system. Based on previous studies, the channel is highly expressed in outer retina, rod inner segments, hippocampus and concentrated in axonal membranes. CONCLUSION: KCNA4 (Kv1.4) is implicated in a novel syndrome characterised by striatal thinning, congenital cataract and attention deficit hyperactivity disorder. Our study highlights potassium channels' role in ocular and neuronal genetics.

ISCA2 mutation causes infantile neurodegenerative mitochondrial disorder.

BACKGROUND: There are numerous nuclear genes that cause mitochondrial disorders and clinically and genetically heterogeneous disorders whose aetiology often remains unsolved. In this study, we aim to investigate an autosomal recessive syndrome causing leukodystrophy and neuroregression. We studied six patients from five unrelated consanguineous families. METHODS: Patients underwent full neurological, radiological, genetic, metabolic and dysmorphological examinations. Exome sequencing coupled with autozygosity mapping, Sanger sequencing, microsatellite haplotyping, standard and molecular karyotyping and whole mitochondrial DNA sequencing were used to identify the genetic cause of the syndrome. Immunohistochemistry, transmission electron microscopy, confocal microscopy, dipstick assays, quantitative PCR, reverse transcription PCR and quantitative reverse transcription PCR were performed on different tissue samples from the patients. RESULTS: We identified a homoallelic missense founder mutation in ISCA2 leading to mitochondrial depletion and reduced complex I activity as well as decreased ISCA2, ISCA1 and IBA57 expression in fibroblasts. MRI indicated similar white matter abnormalities in the patients. Histological examination of the skeletal muscle showed mild to moderate variation in myofibre size and the presence of many randomly distributed atrophic fibres. CONCLUSIONS: Our data demonstrate that ISCA2 deficiency leads to a hereditary mitochondrial neurodegenerative white matter disease in infancy.

Novel homozygous DEAF1 variant suspected in causing white matter disease, intellectual disability, and microcephaly.

DEAF1 encodes a transcriptional binding factor and is a regulator of serotonin receptor 1A. Its protein has a significant expression in the neurons of different brain regions and is involved in early embryonic development. In addition, its role in neural tube development is evident from the knockout mouse as many homozygotes have exencephaly. Heterozygous mutations of this gene have been linked to intellectual disability in addition to the gene's involvement in major depression, suicidal tendencies, and panic disorder. In this clinical report, we describe two children from a consanguineous family with intellectual disability, microcephaly, and hypotonia. The brain MRI of both patients showed bilateral and symmetrical white matter abnormalities, and one of the patients had a seizure disorder. Using whole exome sequencing combined with homozygosity mapping, a homozygous p.R226W (c.676C>T) mutation in DEAF1 was found in both patients. Furthermore, sequencing analysis confirmed complete segregation in tested family members and absence of the mutation in control cohort (n = 650). The mutation is located in a highly conserved structural domain that mediates DNA binding and therefore regulates transcriptional activity of its target molecules. This study indicates, for the first time to our knowledge, a hereditary role of DEAF1 in white matter abnormalities, microcephaly and syndromic intellectual disability.

Mutations in NALCN cause an autosomal-recessive syndrome with severe hypotonia, speech impairment, and cognitive delay.

Sodium leak channel, nonselective (NALCN) is a voltage-independent and cation-nonselective channel that is mainly responsible for the leaky sodium transport across neuronal membranes and controls neuronal excitability. Although NALCN variants have been conflictingly reported to be in linkage disequilibrium with schizophrenia and bipolar disorder, to our knowledge, no mutations have been reported to date for any inherited disorders. Using linkage, SNP-based homozygosity mapping, targeted sequencing, and confirmatory exome sequencing, we identified two mutations, one missense and one nonsense, in NALCN in two unrelated families. The mutations cause an autosomal-recessive syndrome characterized by subtle facial dysmorphism, variable degrees of hypotonia, speech impairment, chronic constipation, and intellectual disability. Furthermore, one of the families pursued preimplantation genetic diagnosis on the basis of the results from this study, and the mother recently delivered healthy twins, a boy and a girl, with no symptoms of hypotonia, which was present in all the affected children at birth. Hence, the two families we describe here represent instances of loss of function in human NALCN.