A Biallelic Variant of the RNA Exosome Gene, EXOSC4, Associated with Neurodevelopmental Defects Impairs RNA Exosome Function and Translation.

The RNA exosome is an evolutionarily conserved complex required for both precise RNA processing and decay. Pathogenic variants in EXOSC genes, which encode structural subunits of this complex, are linked to several autosomal recessive disorders. Here, we describe a missense allele of the EXOSC4 gene that causes a collection of clinical features in two affected siblings. This missense variant (NM_019037.3: exon3:c.560T>C), changes a leucine residue within a conserved region of EXOSC4 to proline (p.Leu187Pro). The two affected individuals show prenatal growth restriction, failure to thrive, global developmental delay, intracerebral and basal ganglia calcifications, and kidney failure. Homozygosity for the damaging variant was identified by exome sequencing with Sanger sequencing to confirm segregation. To explore the functional consequences of this amino acid change, we modeled EXOSC4-L187P in the corresponding budding yeast protein, Rrp41 (Rrp41-L187P). Cells that express Rrp41-L187P as the sole copy of the essential Rrp41 protein show growth defects. Steady-state levels of both Rrp41-L187P and EXOSC4-L187P are decreased compared to controls and EXOSC4-L187P shows decreased co-purification with other RNA exosome subunits. RNA exosome target transcripts accumulate in rrp41-L187P cells, including the 7S precursor of 5.8S rRNA. Polysome profiles show a decrease in actively translating ribosomes in rrp41-L187P cells as compared to control cells with incorporation of 7S pre-rRNA into polysomes. This work adds EXOSC4 to the structural subunits of the RNA exosome that have been linked to human disease and defines foundational molecular defects that could contribute to the adverse phenotypes caused by EXOSC pathogenic variants.

A Biallelic Variant of the RNA Exosome Gene EXOSC4 Causes Translational Defects Associated with a Neurodevelopmental Disorder.

The RNA exosome is an evolutionarily conserved complex required for both precise RNA processing and decay. Mutations in EXOSC genes encoding structural subunits of the complex are linked to several autosomal recessive disorders. Here, we describe a missense allele of the EXOSC4 gene, which causes a collection of clinical features in two affected siblings. This missense mutation (NM_019037.3: exon3:c.560T>C), changes a leucine residue within a highly conserved region of EXOSC4 to proline (p.Leu187Pro). The two affected individuals presented with prenatal growth restriction, failure to thrive, global developmental delay, intracerebral and basal ganglia calcifications, and kidney failure. Homozygosity for the damaging variant was identified through exome sequencing and Sanger sequencing confirmed segregation. To explore the functional consequences of this amino acid change, we modeled EXOSC4-L187P in the corresponding budding yeast protein, Rrp41 (Rrp41-L187P). Cells that express Rrp41-L187P as the sole copy of the essential Rrp41 protein show significant growth defects. The steady-state level of both the Rrp41-L187P and the EXOSC4-L187P proteins is significantly decreased compared to control Rrp41/EXOSC4. Consistent with this observation, targets of the RNA exosome accumulate in rrp41-L187P cells, including the 7S precursor of 5.8S rRNA. Polysome profiles show a significant decrease in translation in rrp41-L187P cells as compared to control cells with apparent incorporation of 7S pre-rRNA into polysomes. Taken together, this work adds the EXOSC4 subunit of the RNA exosome to the structural subunits of this complex that have been linked to human disease and defines foundational molecular defects that could contribute to the adverse growth phenotypes caused by this novel EXOSC4 pathogenic variant.

The super sickling haemoglobin HbS-Oman: a study of red cell sickling, K+ permeability and associations with disease severity in patients heterozygous for HbA and HbS-Oman (HbA/S-Oman genotype).

Studying different sickle cell genotypes may throw light on the pathogenesis of sickle cell disease (SCD). Here, the clinical profile, red cell sickling and K+ permeability in 29 SCD patients (15 patients with severe disease and 14 with a milder form) of HbA/S-Oman genotype were analysed. The super sickling nature of this Hb variant was confirmed. The red cell membrane permeability to K+ was markedly abnormal with elevated activities of Psickle , Gardos channel and KCl cotransporter (KCC). Results were consistent with Ca2+ entry and Mg2+ loss via Psickle stimulating Gardos channel and KCC activities. The abnormal red cell behaviour was similar to that in the commonest genotype of SCD, HbSS, in which the level of mutated Hb is considerably higher. Although activities of all three K+ transporters also correlated with the level of HbS-Oman, there was no association between transport phenotype and disease severity. The super sickling behaviour of HbS-Oman may obviate the need for solute loss and red cell dehydration to encourage Hb polymerisation, required in other SCD genotypes. Disease severity was reduced by concurrent α thalassaemia, as observed in other SCD genotypes, and represents an obvious genetic marker for prognostic tests of severity in young SCD patients of the HbA/S-Oman genotype.

A novel mutation in DDR2 causing spondylo-meta-epiphyseal dysplasia with short limbs and abnormal calcifications (SMED-SL) results in defective intra-cellular trafficking.

BACKGROUND: The rare autosomal genetic disorder, Spondylo-meta-epiphyseal dysplasia with short limbs and abnormal calcifications (SMED-SL), is reported to be caused by missense or splice site mutations in the human discoidin domain receptor 2 (DDR2) gene. Previously our group has established that trafficking defects and loss of ligand binding are the underlying cellular mechanisms of several SMED-SL causing mutations. Here we report the clinical characteristics of two siblings of consanguineous marriage with suspected SMED-SL and identification of a novel disease-causing mutation in the DDR2 gene. METHODS: Clinical evaluation and radiography were performed to evaluate the patients. All the coding exons and splice sites of the DDR2 gene were sequenced by Sanger sequencing. Subcellular localization of the mutated DDR2 protein was determined by confocal microscopy, deglycosylation assay and Western blotting. DDR2 activity was measured by collagen activation and Western analysis. RESULTS: In addition to the typical features of SMED-SL, one of the patients has an eye phenotype including visual impairment due to optic atrophy. DNA sequencing revealed a novel homozygous dinucleotide deletion mutation (c.2468_2469delCT) on exon 18 of the DDR2 gene in both patients. The mutation resulted in a frameshift leading to an amino acid change at position S823 and a predicted premature termination of translation (p.S823Cfs*2). Subcellular localization of the mutant protein was analyzed in mammalian cell lines, and it was found to be largely retained in the endoplasmic reticulum (ER), which was further supported by its N-glycosylation profile. In keeping with its cellular mis-localization, the mutant protein was found to be deficient in collagen-induced receptor activation, suggesting protein trafficking defects as the major cellular mechanism underlying the loss of DDR2 function in our patients. CONCLUSIONS: Our results indicate that the novel mutation results in defective trafficking of the DDR2 protein leading to loss of function and disease. This confirms our previous findings that DDR2 missense mutations occurring at the kinase domain result in retention of the mutant protein in the ER.

Transglutaminase-1 mutations in Omani families with lamellar ichthyosis.

OBJECTIVE: To determine the molecular basis of familial ichthyosis in three Omani families. SUBJECTS AND METHODS: Nine patients from three consanguineous families, A, B, and C, were born with typical features of lamellar ichthyosis subtype including collodion membrane and maintained ectropion, and epidermal scaling through their childhood. The 4 patients from family B had more severe symptoms requiring neonatal critical care and subsequent regular treatment with emollients, eye lubricants, and low-dose acitretin. DNA was extracted from peripheral blood by standard methods. The samples were initially genotyped to screen known loci linked to recessive ichthyosis on chromosomes 2q33-32 (ABCA12), 14q11 (TGM1), and 19p12-q12 using commercially supplied polymorphic fluorescent microsatellite markers. TGM1 was analyzed by direct sequencing for disease-associated mutations. RESULTS: Two known pathogenic mutations in TGM1 were detected: p.Gly278Arg in families A and B and p.Arg396His in family C. These two mutations were segregating in an autosomal recessive mode of inheritance. CONCLUSION: Two known pathogenic TGM1 mutations were detected in three large consanguineous Omani families with lamellar ichthyosis. This study confirmed the geographic distribution of known mutations to an apparently unrelated population.