A Limited Structural Modification Results in a Significantly More Efficacious Diazachrysene-Based Filovirus Inhibitor

Ebola (EBOV) and Marburg (MARV) filoviruses are highly infectious pathogens causing deadly hemorrhagic fever in humans and non-human primates. Promising vaccine candidates providing immunity against filoviruses have been reported. However, the sporadic nature and swift progression of filovirus disease underlines the need for the development of small molecule therapeutics providing immediate antiviral effects. Herein we describe a brief structural exploration of two previously reported diazachrysene (DAAC)-based EBOV inhibitors. Specifically, three analogs were prepared to examine how slight substituent modifications would affect inhibitory efficacy and inhibitor-mediated toxicity during not only EBOV, but also MARV cellular infection. Of the three analogs, one was highly efficacious, providing IC50 values of 0.696 mu M +/- 0.13 mu M and 2.76 mu M +/- 0.21 mu M against EBOV and MARV infection, respectively, with little or no associated cellular toxicity. Overall, the structure-activity and structure-toxicity results from this study provide a framework for the future development of DAAC-based filovirus inhibitors that will be both active and non-toxic in vivo

Biallelic Mutations in MRPS34 Lead to Instability of the Small Mitoribosomal Subunit and Leigh Syndrome

The synthesis of all 13 mitochondrial DNA (mtDNA)-encoded protein subunits of the human oxidative phosphorylation (OXPHOS) system is carried out by mitochondrial ribosomes (mitoribosomes). Defects in the stability of mitoribosomal proteins or mitoribosome assembly impair mitochondrial protein translation, causing combined OXPHOS enzyme deficiency and clinical disease. Here we report four autosomal-recessive pathogenic mutations in the gene encoding the small mitoribosomal subunit protein, MRPS34, in six subjects from four unrelated families with Leigh syndrome and combined OXPHOS defects. Whole-exome sequencing was used to independently identify all variants. Two splice-site mutations were identified, including homozygous c.321+1G>T in a subject of Italian ancestry and homozygous c.322−10G>A in affected sibling pairs from two unrelated families of Puerto Rican descent. In addition, compound heterozygous MRPS34 mutations were identified in a proband of French ancestry; a missense (c.37G>A [p.Glu13Lys]) and a nonsense (c.94C>T [p.Gln32∗]) variant. We demonstrated that these mutations reduce MRPS34 protein levels and the synthesis of OXPHOS subunits encoded by mtDNA. Examination of the mitoribosome profile and quantitative proteomics showed that the mitochondrial translation defect was caused by destabilization of the small mitoribosomal subunit and impaired monosome assembly. Lentiviral-mediated expression of wild-type MRPS34 rescued the defect in mitochondrial translation observed in skin fibroblasts from affected subjects, confirming the pathogenicity of MRPS34 mutations. Our data establish that MRPS34 is required for normal function of the mitoribosome in humans and furthermore demonstrate the power of quantitative proteomic analysis to identify signatures of defects in specific cellular pathways in fibroblasts from subjects with inherited disease

Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

The complete sequence of human chromosome 5

Chromosome 5 is one of the largest human chromosomes yet has one of the lowest gene densities. This is partially explained by numerous gene-poor regions that display a remarkable degree of noncoding and syntenic conservation with non-mammalian vertebrates, suggesting they are functionally constrained. In total, we compiled 177.7 million base pairs of highly accurate finished sequence containing 923 manually curated protein-encoding genes including the protocadherin and interleukin gene families and the first complete versions of each of the large chromosome 5 specific internal duplications. These duplications are very recent evolutionary events and play a likely mechanistic role, since deletions of these regions are the cause of debilitating disorders including spinal muscular atrophy (SMA)

Post-Intoxication Inhibition of Botulinum Neurotoxin Serotype A within Neurons by Small-Molecule, Non-Peptidic Inhibitors

Botulinum neurotoxins (BoNTs) comprise seven distinct serotypes that inhibit the release of neurotransmitter across neuromuscular junctions, resulting in potentially fatal flaccid paralysis. BoNT serotype A (BoNT/A), which targets synaptosomal-associated protein of 25kDa (SNAP-25), is particularly long-lived within neurons and requires a longer time for recovery of neuromuscular function. There are currently no treatments available to counteract BoNT/A after it has entered the neuronal cytosol. In this study, we examined the ability of small molecule non-peptidic inhibitors (SMNPIs) to prevent SNAP-25 cleavage post-intoxication of neurons. The progressive cleavage of SNAP-25 observed over 5 h following 1 h BoNT/A intoxication was prevented by addition of SMNPIs. In contrast, anti-BoNT/A neutralizing antibodies that strongly inhibited SNAP-25 cleavage when added during intoxication were completely ineffective when added post-intoxication. Although Bafilomycin A1, which blocks entry of BoNT/A into the cytosol by preventing endosomal acidification, inhibited SNAP-25 cleavage post-intoxication, the degree of inhibition was significantly reduced versus addition both during and after intoxication. Post-intoxication application of SMNPIs, on the other hand, was nearly as effective as application both during and after intoxication. Taken together, the results indicate that competitive SMNPIs of BoNT/A light chain can be effective within neurons post-intoxication

α-Gal specific-IgE prevalence and levels in Ecuador and Kenya: Relation to diet, parasites, and IgG4

Background IgE to α-Gal is a cause of mammalian meat allergy and has been linked to tick bites in North America, Australia, and Eurasia. Reports from the developing world indicate that α-Gal sensitization is prevalent but has been little investigated. Objective We sought evidence for the cause(s) of α-Gal sensitization and lack of reported meat allergy among children in less developed settings in Ecuador and Kenya. Methods IgE to α-Gal and total IgE were assessed in children from Ecuador (n = 599) and Kenya (n = 254) and compared with children with (n = 42) and without known (n = 63) mammalian meat allergy from the southeastern United States. Information on diet, potential risk factors, and helminth infections was available for children from Ecuador. IgG4 to α-Gal and antibodies to regionally representative parasites were assessed in a subset of children. Results In Ecuador (32%) and Kenya (54%), α-Gal specific IgE was prevalent, but levels were lower than in children with meat allergy from the United States. Sensitization was associated with rural living, antibody markers of Ascaris exposure, and total IgE, but not active infections with Ascaris or Trichuris species. In Ecuador, 87.5% reported consuming beef at least once per week, including 83.9% of those who had α-Gal specific IgE. Levels of α-Gal specific IgG4 were not high in Ecuador, but were greater than in children from the United States. Conclusions These results suggest that in areas of the developing world with endemic parasitism, α-Gal sensitization is (1) common, (2) associated with Ascaris exposure, and (3) distinguished by a low percentage of specific/total IgE compared with individuals with meat allergy in the United States

Scintillation Observations and Response of the Ionoshere to Electrodynamics (SORTIE)

At low and middle latitudes, wavelike plasma perturbations are thought to provide the seeds for larger perturbations that may evolve non-linearly to produce irregularities which in turn have deleterious effects on HF communications and global positioning systems. However, there is currently no comprehensive atlas of measurements describing the global spatial or temporal distribution of wave-like perturbations in the ionosphere. The SORTIE mission is a 6U CubeSat mission with team members from ASTRA, AFRL, UTD, COSMIAC, and Boston College. The SORTIE spacecraft is designed to approach the complex challenges in discovering the wave-like plasma perturbations in the ionosphere. SORTIE will provide the initial spectrum of wave perturbations which are the starting point for the RF calculation, provide measured electric fields which determine the magnitude of the instability growth rate near where plasma bubbles are generated, and will provide initial observations of the irregularities in plasma density which result from instability growth. The SORTIE mission is slated to launch in late 2017, and will provide a timely overlap with NASA\u27s ICON mission scheduled to launch in the 2017 timeframe. The baseline operational plan will be a year of on-orbit lifetime orbiting at a low to middle inclination orbit near 350-400 km altitude