Exome sequencing identifies ACAD9 mutations as a cause of complex I deficiency

An isolated defect of respiratory chain complex I activity is a frequent biochemical abnormality in mitochondrial disorders. Despite intensive investigation in recent years, in most instances, the molecular basis underpinning complex I defects remains unknown. We report whole-exome sequencing of a single individual with severe, isolated complex I deficiency. This analysis, followed by filtering with a prioritization of mitochondrial proteins, led us to identify compound heterozygous mutations in ACAD9, which encodes a poorly understood member of the mitochondrial acyl-CoA dehydrogenase protein family. We demonstrated the pathogenic role of the ACAD9 variants by the correction of the complex I defect on expression of the wildtype ACAD9 protein in fibroblasts derived from affected individuals. ACAD9 screening of 120 additional complex I-defective index cases led us to identify two additional unrelated cases and a total of five pathogenic ACAD9 alleles.     

Osteoclast-poor human osteopetrosis due to mutations in the gene encoding RANKL

Autosomal recessive osteopetrosis is usually associated with normal or elevated numbers of nonfunctional osteoclasts. Here we report mutations in the gene encoding RANKL (receptor activator of nuclear factor-KB ligand) in six individuals with autosomal recessive osteopetrosis whose bone biopsy specimens lacked osteoclasts. These individuals did not show any obvious defects in immunological parameters and could not be cured by hematopoietic stem cell transplantation; however, exogenous RANKL induced formation of functional osteoclasts from their monocytes, suggesting that they could, theoretically, benefit from exogenous RANKL administration.     

Comparative myogenesis in teleosts and mammals

Skeletal myogenesis has been and is currently under extensive study in both mammals and teleosts, with the latter providing a good model for skeletal myogenesis because of their flexible and conserved genome. Parallel investigations of muscle studies using both these models have strongly accelerated the advances in the field. However, when transferring the knowledge from one model to the other, it is important to take into account both their similarities and differences. The main difficulties in comparing mammals and teleosts arise from their different temporal development. Conserved aspects can be seen for muscle developmental origin and segmentation, and for the presence of multiple myogenic waves. Among the divergences, many fish have an indeterminate growth capacity throughout their entire life span, which is absent in mammals, thus implying different post-natal growth mechanisms. This review covers the current state of the art on myogenesis, with a focus on the most conserved and divergent aspects between mammals and teleosts.     

The positional identity of mouse ES cell-generated neurons is affected by BMP signaling

We investigated the effects of bone morphogenetic proteins (BMPs) in determining the positional identity of neurons generated in vitro from mouse embryonic stem cells (ESCs), an aspect that has been neglected thus far. Classical embryological studies in lower vertebrates indicate that BMPs inhibit the default fate of pluripotent embryonic cells, which is both neural and anterior. Moreover, mammalian ESCs generate neurons more efficiently when cultured in a minimal medium containing BMP inhibitors. In this paper, we show that mouse ESCs produce, secrete, and respond to BMPs during in vitro neural differentiation. After neuralization in a minimal medium, differentiated ESCs show a gene expression profile consistent with a midbrain identity, as evaluated by the analysis of a number of markers of anterior–posterior and dorsoventral identity. We found that BMPs endogenously produced during neural differentiation mainly act by inhibiting the expression of a telencephalic gene profile, which was revealed by the treatment with Noggin or with other BMP inhibitors. To better characterize the effect of BMPs on positional fate, we compared the global gene expression profiles of differentiated ESCs with those of embryonic forebrain, midbrain, and hindbrain. Both Noggin and retinoic acid (RA) support neuronal differentiation of ESCs, but they show different effects on their positional identity: whereas RA supports the typical gene expression profile of hindbrain neurons, Noggin induces a profile characteristic of dorsal telencephalic neurons. Our findings show that endogenously produced BMPs affect the positional identity of the neurons that ESCs spontaneously generate when differentiating in vitro in a minimal medium. The data also support the existence of an intrinsic program of neuronal differentiation with dorsal telencephalic identity. Our method of ESC neuralization allows for fast differentiation of neural cells via the same signals found during in vivo embryonic development and for the acquisition of cortical identity by the inhibition of BMP alone.     

Mutations in TTC19 cause mitochondrial complex III deficiency and neurological impairment in humans and flies

Although mutations in CYTB (cytochrome b) or BCS1L have been reported in isolated defects of mitochondrial respiratory chain complex III (cIII), most cIII-defective individuals remain genetically undefined. We identified a homozygous nonsense mutation in the gene encoding tetratricopeptide 19 (TTC19) in individuals from two families affected by progressive encephalopathy associated with profound cIII deficiency and accumulation of cIII-specific assembly intermediates. We later found a second homozygous nonsense mutation in a fourth affected individual. We demonstrated that TTC19 is embedded in the inner mitochondrial membrane as part of two high-molecular-weight complexes, one of which coincides with cIII. We then showed a physical interaction between TTC19 and cIII by coimmunoprecipitation. We also investigated a Drosophila melanogaster knockout model for TTC19 that showed low fertility, adult-onset locomotor impairment and bang sensitivity, associated with cIII deficiency. TTC19 is a putative cIII assembly factor whose disruption is associated with severe neurological abnormalities in humans and flies.     

Analysis of the coding genome of diffuse large B-cell lymphoma

Diffuse large B-cell lymphoma (DLBCL) is the most common form of human lymphoma. Although a number of structural alterations have been associated with the pathogenesis of this malignancy, the full spectrum of genetic lesions that are present in the DLBCL genome, and therefore the identity of dysregulated cellular pathways, remains unknown. By combining next-generation sequencing and copy number analysis, we show that the DLBCL coding genome contains, on average, more than 30 clonally represented gene alterations per case. This analysis also revealed mutations in genes not previously implicated in DLBCL pathogenesis, including those regulating chromatin methylation (MLL2; 24% of samples) and immune recognition by T cells. These results provide initial data on the complexity of the DLBCL coding genome and identify novel dysregulated pathways underlying its pathogenesis.     

Killing activity of neutrophils is mediated through activation of proteases by K+ flux

According to the hitherto accepted view, neutrophils kill ingested microorganisms by subjecting them to high concentrations of highly toxic reactive oxygen species (ROS) and bringing about myeloperoxidase-catalysed halogenation. We show here that this simple scheme, which for many years has served as a satisfactory working hypothesis, is inadequate. We find that mice made deficient in neutrophil-granule proteases but normal in respect of superoxide production and iodinating capacity, are unable to resist staphylococcal and candidal infections. We also show that activation provokes the influx of an enormous concentration of ROS into the endocytic vacuole. The resulting accumulation of anionic charge is compensated for by a surge of K+ ions that cross the membrane in a pH-dependent manner. The consequent rise in ionic strength engenders the release of cationic granule proteins, including elastase and cathepsin G, from the anionic sulphated proteoglycan matrix. We show that it is the proteases, thus activated, that are primarily responsible for the destruction of the bacteria.     

A spectrophotometric method for the determination of 5-phosphoribosyl-1-pyrophosphate

Summary A new, sensitive, specific and simple spectrophotometric method for the determination of 5-phosphoribosyl-1-pyrophosphate (PRPP) is presented. PRPP is reacted with excess hypoxanthine in the presence of hypoxanthine-guanine phosphoribosyltransferase. At the end of the reaction, PRPP concentration is measured from the extent of conversion of hypoxanthine to inositate. The concentration of the purine base is determined spectrophotometrically in the presence of xanthine oxidase.     

Mitochondrial aspartyl-tRNA synthetase deficiency causes leukoencephalopathy with brain stem and spinal cord involvement and lactate elevation

Leukoencephalopathy with brain stem and spinal cord involvement and lactate elevation (LBSL) has recently been defined based on a highly characteristic constellation of abnormalities observed by magnetic resonance imaging and spectroscopy. LBSL is an autosomal recessive disease, most often manifesting in early childhood. Affected individuals develop slowly progressive cerebellar ataxia, spasticity and dorsal column dysfunction, sometimes with a mild cognitive deficit or decline. We performed linkage mapping with microsatellite markers in LBSL families and found a candidate region on chromosome 1, which we narrowed by means of shared haplotypes. Sequencing of genes in this candidate region uncovered mutations in DARS2, which encodes mitochondrial aspartyl-tRNA synthetase, in affected individuals from all 30 families. Enzyme activities of mutant proteins were decreased. We were surprised to find that activities of mitochondrial complexes from fibroblasts and lymphoblasts derived from affected individuals were normal, as determined by different assays.     

Deficiency of hyccin, a newly identified membrane protein, causes hypomyelination and congenital cataract

We describe a new autosomal recessive white matter disorder ('hypomyelination and congenital cataract') characterized by hypomyelination of the central and peripheral nervous system, progressive neurological impairment and congenital cataract. We identified mutations in five affected families, resulting in a deficiency of hyccin, a newly identified 521-amino acid membrane protein. Our study highlights the essential role of hyccin in central and peripheral myelination.