Genome sequencing in persistently unsolved white matter disorders

Genetic white matter disorders have heterogeneous etiologies and overlapping clinical presentations. We performed a study of the diagnostic efficacy of genome sequencing in 41 unsolved cases with prior exome sequencing, resolving an additional 14 from an historical cohort (n = 191). Reanalysis in the context of novel disease-associated genes and improved variant curation and annotation resolved 64% of cases. The remaining diagnoses were directly attributable to genome sequencing, including cases with small and large copy number variants (CNVs) and variants in deep intronic and technically difficult regions. Genome sequencing, in combination with other methodologies, achieved a diagnostic yield of 85% in this retrospective cohort.Guy Helman, Bryan R. Lajoie, Joanna Crawford, Asako Takanohashi, Marzena Walkiewicz ... Stephen J. Bent ... et al

Identification of RNA helicases in human immunodeficiency virus 1 (HIV-1) replication – A targeted small interfering RNA library screen using pseudotyped and WT HIV-1

10.1099/vir.0.000092Journal of General Virology961484-148

Evidence that the endosomal sorting complex required for transport-II (ESCRT-II) is required for efficient human immunodeficiency virus-1 (HIV-1) production

10.1186/s12977-015-0197-xRetrovirology1217

Eight novel mutations in MLC1 from 18 Iranian patients with megalencephalic leukoencephalopathy with subcortical cysts

Megalencephalic leukoencephalopathy with subcortical cysts (MLC) (MIM #. 604004) is a rare autosomal recessive neurological disorder characterized by macrocephaly, motor and cognitive decline, ataxia, spasticity and occasional seizures. Magnetic resonance imaging (MRI) shows diffusely abnormal and swollen white matter of the cerebral hemispheres and subcortical cysts in the anterior temporal and frontoparietal region. Mutations in MLC1(22q13.33) and GLIALCAM have been identified in patients with MLC. Mutations in MLC1 account for approximately 75 of the cases.MLC was suspected in eighteen Iranian patients from sixteen families based on positive clinical findings including macrocephaly beginning in the first year, neurocognitive deterioration, seizure or loss of consciousness after minor head trauma. All except two were born to consanguineous parents. Brain MRI images were compatible with MLC and confirmed the diagnosis. Sequencing of entire coding region of MLC1 was performed for seventeen patients and mutations in MLC1 were detected in all of them. Eight novel mutations and seven previously reported mutations were identified. This report shows that MLC is relatively common in Iranian population, as expected for rare diseases with high inbreeding, with a surprisingly high frequency of novel mutations. © 2014 Elsevier Masson SAS

Mice with megalencephalic leukoencephalopathy with cysts: a developmental angle

Objective: Megalencephalic leukoencephalopathy with cysts (MLC) is a genetic disease characterized by infantile onset white matter edema and delayed onset neurological deterioration. Loss of MLC1 function causes MLC. MLC1 is involved in ion-water homeostasis, but its exact role is unknown. We generated Mlc1-null mice for further studies. Methods: We investigated which brain cell types express MLC1, compared developmental expression in mice and men, and studied the consequences of loss of MLC1 in Mlc1-null mice. Results: Like humans, mice expressed MLC1 only in astrocytes, especially those facing fluid-brain barriers. In mice, MLC1 expression increased until 3 weeks and then stabilized. In humans, MLC1 expression was highest in the first year, decreased, and stabilized from approximately 5 years. Mlc1-null mice had early onset megalencephaly and increased brain water content. From 3 weeks, abnormal astrocytes were present with swollen processes abutting fluid-brain barriers. From 3 months, widespread white matter vacuolization with intramyelinic edema developed. Mlc1-null astrocytes showed slowed regulatory volume decrease and reduced volume-regulated anion currents, which increased upon MLC1 re-expression. Mlc1-null astrocytes showed reduced expression of adhesion molecule GlialCAM and chloride channel ClC-2, but no substantial changes in other known MLC1-interacting proteins. Interpretation: Mlc1-null mice replicate early stages of the human disease with early onset intramyelinic edema. The cellular functional defects, described for human MLC, were confirmed. The earliest change was astrocytic swelling, substantiating that in MLC the primary defect is in volume regulation by astrocytes. MLC1 expression affects expression of GlialCAM and ClC-2. Abnormal interplay between these proteins is part of the pathomechanisms of MLC

Leukoencephalopathy with brainstem and spinal cord involvement and lactate elevation: clinical and genetic characterization and target for therapy

Leukoencephalopathy with brainstem and spinal cord involvement and lactate elevation is a disorder caused by recessive mutations in the gene DARS2, which encodes mitochondrial aspartyl-tRNA synthetase. Recent observations indicate that the phenotypic range of the disease is much wider than initially thought. Currently, no treatment is available. The aims of our study were (i) to explore a possible genotype-phenotype correlation; and (ii) to identify potential therapeutic agents that modulate the splice site mutations in intron 2 of DARS2, present in almost all patients. A cross-sectional observational study was performed in 78 patients with two DARS2 mutations in the Amsterdam and Helsinki databases up to December 2012. Clinical information was collected via questionnaires. An inventory was made of the DARS2 mutations in these patients and those previously published. An assay was developed to assess mitochondrial aspartyl-tRNA synthetase enzyme activity in cells. Using a fluorescence reporter system we screened for drugs that modulate DARS2 splicing. Clinical information of 66 patients was obtained. The clinical severity varied from infantile onset, rapidly fatal disease to adult onset, slow and mild disease. The most common phenotype was characterized by childhood onset and slow neurological deterioration. Full wheelchair dependency was rare and usually began in adulthood. In total, 60 different DARS2 mutations were identified, 13 of which have not been reported before. Except for 4 of 42 cases published by others, all patients were compound heterozygous. Ninety-four per cent of the patients had a splice site mutation in intron 2. The groups of patients sharing the same two mutations were too small for formal assessment of genotype-phenotype correlation. However, some combinations of mutations were consistently associated with a mild phenotype. The mitochondrial aspartyl-tRNA synthetase activity was strongly reduced in patient cells. Among the compounds screened, cantharidin was identified as the most potent modulator of DARS2 splicing. In conclusion, the phenotypic spectrum of leukoencephalopathy with brainstem and spinal cord involvement and lactate elevation is wide, but most often the disease has a relatively slow and mild course. The available evidence suggests that the genotype influences the phenotype, but because of the high number of private mutations, larger numbers of patients are necessary to confirm this. The activity of mitochondrial aspartyl-tRNA synthetase is significantly reduced in patient cells. A compound screen established a 'proof of principle' that the splice site mutation can be influenced. This finding is promising for future therapeutic strategies. © 2013 The Author