NONO enhances mRNA processing of super‐enhancer‐associated GATA2 and HAND2 genes in neuroblastoma

High‐risk neuroblastoma patients have poor survival rates and require better therapeutic options. High expression of a multifunctional DNA and RNA‐binding protein, NONO, in neuroblastoma is associated with poor patient outcome; however, there is little understanding of the mechanism of NONO‐dependent oncogenic gene regulatory activity in neuroblastoma. Here, we used cell imaging, biochemical and genome‐wide molecular analysis to reveal complex NONO‐dependent regulation of gene expression. NONO forms RNA‐ and DNA‐tethered condensates throughout the nucleus and undergoes phase separation in vitro, modulated by nucleic acid binding. CLIP analyses show that NONO mainly binds to the 5′ end of pre‐mRNAs and modulates pre‐mRNA processing, dependent on its RNA‐binding activity. NONO regulates super‐enhancer‐associated genes, including HAND2 and GATA2. Abrogating NONO RNA binding, or phase separation activity, results in decreased expression of HAND2 and GATA2. Thus, future development of agents that target RNA‐binding activity of NONO may have therapeutic potential in this cancer context.     

Inferring mechanisms of copy number change from haplotype structures at the human DEFA1A3 locus

Background

The determination of structural haplotypes at copy number variable regions can indicate the mechanisms responsible for changes in copy number, as well as explain the relationship between gene copy number and expression. However, obtaining spatial information at regions displaying extensive copy number variation, such as the DEFA1A3 locus, is complex, because of the difficulty in the phasing and assembly of these regions. The DEFA1A3 locus is intriguing in that it falls within a region of high linkage disequilibrium, despite its high variability in copy number (n = 3–16); hence, the mechanisms responsible for changes in copy number at this locus are unclear.

Results

In this study, a region flanking the DEFA1A3 locus was sequenced across 120 independent haplotypes with European ancestry, identifying five common classes of DEFA1A3 haplotype. Assigning DEFA1A3 class to haplotypes within the 1000 Genomes project highlights a significant difference in DEFA1A3 class frequencies between populations with different ancestry. The features of each DEFA1A3 class, for example, the associated DEFA1A3 copy numbers, were initially assessed in a European cohort (n = 599) and replicated in the 1000 Genomes samples, showing within-class similarity, but between-class and between-population differences in the features of the DEFA1A3 locus. Emulsion haplotype fusion-PCR was used to generate 61 structural haplotypes at the DEFA1A3 locus, showing a high within-class similarity in structure.

Conclusions

Structural haplotypes across the DEFA1A3 locus indicate that intra-allelic rearrangement is the predominant mechanism responsible for changes in DEFA1A3 copy number, explaining the conservation of linkage disequilibrium across the locus. The identification of common structural haplotypes at the DEFA1A3 locus could aid studies into how DEFA1A3 copy number influences expression, which is currently unclear.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-614) contains supplementary material, which is available to authorized users.     

Size scaling of whole‐body metabolic activity in the noble crayfish (Astacus astacus) estimated from measurements on a single leg

1. Use of electron transport system (ETS) activity in a single leg for estimating whole‐body ETS activity was explored in the noble crayfish Astacus astacus. Oxygen consumption and ETS activity of the whole body and of a walking leg were measured in different‐sized animals at 10 °C to compare the size scaling of oxygen consumption, whole‐body ETS activity and the ratio of whole‐body ETS activity to oxygen consumption (ETS/R). 2. Electron transport system activity of a leg and the ratio of ETS activity of a whole crayfish to that of a leg were correlated with wet mass of animals. Therefore, metabolic potential in whole noble crayfish can be estimated on the basis of the measured ETS activity in a single leg and crayfish mass. This approach provides a valuable tool for determining metabolic characteristics of crayfish without killing them. 3. Mass‐specific oxygen consumption decreased with increasing wet mass, while ETS activity of whole crayfish showed no significant correlation with wet mass. Both oxygen consumption and ETS activity correlated significantly with protein mass. 4. The increase in ETS/R with increasing wet mass of the noble crayfish indicates that small organisms exploit a greater proportion of their metabolic potential for standard metabolism than larger ones. This is the first report on ETS/R in crayfish.     

Production of functionally active <Emphasis Type="Italic">Penicillium chrysogenum</Emphasis> isopenicillin N synthase in the yeast <Emphasis Type="Italic">Hansenula polymorpha</Emphasis>

Background  

β-Lactams like penicillin and cephalosporin are among the oldest known antibiotics used against bacterial infections. Industrially, penicillin is produced by the filamentous fungus Penicillium chrysogenum. Our goal is to introduce the entire penicillin biosynthesis pathway into the methylotrophic yeast Hansenula polymorpha. Yeast species have the advantage of being versatile, easy to handle and cultivate, and possess superior fermentation properties relative to filamentous fungi. One of the fundamental challenges is to produce functionally active enzyme in H. polymorpha.     

AN EARLY CENOZOIC NEOSELACHIAN SHARK FAUNA FROM THE SOUTHWEST PACIFIC

Abstract: An early Cenozoic shark fauna, comprising at least 16 taxa, is described from Paleocene sedimentary rocks on the South Island of New Zealand. Although representing a remote Southern Hemisphere location, the fauna includes forms closely comparable to contemporary species from the Northern Hemisphere, in addition to the new species Chlamydoselachus keyesi and Centroselachus goordi. Comparison with closely related extant species suggests the fauna may be interpreted as a deep water one, typical of the outer continental shelf and upper slope. However, after palaeogeography, sedimentology and mineralogy of the enclosing rock, and the nature of similar faunas from elsewhere are taken into consideration, the fauna is interpreted to have occupied a mid‐shelf environment.     

Hybridization between Crocodylus acutus and Crocodylus moreletii in the Yucatan Peninsula: I. Evidence from mitochondrial DNA and morphology

The American crocodile (Crocodylus acutus) and the Morelet's crocodile (C. moreletii) are broadly sympatric in Belize and Mexico. The presence of morphologically anomalous individuals in the overlapping range area suggests possible hybridization between these species. Analysis of 477 base pairs of the mitochondrial tRNA(Pro)-tRNA(Phe)-Dloop region revealed the presence of pure C. acutus (N=43) and C. moreletii (N=56), as well as a high proportion of interspecific hybrids (N=17, 14.6%) in the Yucatan Peninsula, Mexico. Although all individuals could be assigned to one species or other based on phenotypic characters, some had been characterized as potential hybrids in the field by anomalous scale counts. The hybridization zone lies along the area of sympatry between C. acutus and C. moreletii investigated in this study, but extends further inland if hybrid localities from Belize are included. Hybridization in the Yucatan Peninsula is bidirectional, which indicates considerably more genetic contact between these species than previously recognized, and is probably more detrimental to the genetic integrity of smaller C. acutus populations. A more intensive study of the pattern of hybridization is warranted and supports continued classification of C. acutus as a critically threatened species in the Yucatan Peninsula.     

Low Levels of Nucleotide Diversity in Crocodylus moreletii and Evidence of Hybridization with C. acutus

Examinations of both population genetic structure and the processes that lead to such structure in crocodilians have been initiated in several species in response to a call by the IUCN Crocodile Specialist Group. A recent study used microsatellite markers to characterize Morelet's crocodile (Crocodylus moreletii) populations in north-central Belize and presented evidence for isolation by distance. To further investigate this hypothesis, we sequenced a portion of the mitochondrial control region for representative animals after including samples from additional locales in Belize, Guatemala and Mexico. While there is limited evidence of subdivision involving other locales, we found that most of the differentiation among populations of C. moreletiican be attributed to animals collected from a single locale in Belize, Banana Bank Lagoon. Furthermore, mitochondrial DNA sequence analysis showed that animals from this and certain other locales display a haplotype characteristic of the American crocodile, C. acutus, rather than C. moreletii. We interpret this as evidence of hybridization between the two species and comment on how these new data have influenced our interpretation of previous findings. We also find very low levels of nucleotide diversity in C. moreletiihaplotypes and provide evidence for a low rate of substitution in the crocodilian mitochondrial control region. Finally, the conservation implications of these findings are discussed.     

Effects of External Mass Transfer and Product Inhibition on a Simulated Immobilized Enzyme‐Catalyzed Reactor for Lactose Hydrolysis

A mathematical model has been developed for predicting the performance and simulation of a packed bed immobilized enzyme reactor performing lactose hydrolysis, which follows Michaelis‐Menten kinetics with competitive product (galactose) inhibition. The performance characteristics of a packed bed immobilized enzyme reactor have been analyzed taking into account the effects of various diffusional phenomena like axial dispersion and external mass transfer limitations. The model design equations are then solved by Galerkin's method and orthogonal collocation on finite elements. The effects of external mass transfer and axial dispersion have been studied and their effects were shown to reduce the external effectiveness factor. The effects of product inhibition have been investigated at different operating conditions correlated at different regimes using dimensionless moduli (St, γ, θ, Da)¹⁾. The product inhibition was shown to reduce the substrate conversion, and, additionally, to decrease the effectiveness factor when Da > Da_xo, however, it increases the effectiveness factor when Da < Da_xo. The effectiveness factor is found to be independent of the product inhibition at a crossover point at which Da_xo is defined. Effects of St and Pe have been investigated at different kinetic regimes and the results show that their effects have a strong dependency on the kinetic parameters θ, γ (i.e., K_m/K_p), and Da_xo.     

在中国应用垂直波束雷达监测褐飞虱和其他水稻害虫迁飞的可行性

近年来,可用于昆虫迁飞研究且可自动运行的垂直波束雷达(vertical-looking radar,VLR)的发展使得对迁飞性害虫的周年长期自动监测成为可能。本文提供了我们对能否将这种雷达应用于中国的褐飞虱和其他水稻害虫的监测与预测体系以改善其综合治理的可行性研究结果。以往的研究已经表明,这些害虫一般在300—2000m高度迁飞;而我们根据褐飞虱的雷达和射有效截面的计算结果表明,目前使用的3．2cm波长的VLR对褐飞虱个体目标的最大可检测高度仅约240m;虽然建造一部8．8mm波长的VLR即可覆盖褐飞虱迁飞高度的绝大部分,但其造价和维护费用均过于昂贵。为此,一个更可行的解决方案是,以3．2cm波长的VLR作为包括大多数水稻害虫在内的个体较大的迁飞性害虫的监测工具。     

Protection against influenza infection by cytokine-enhanced aerosol genetic immunization

BACKGROUND: Conventional vaccine development for newly emerging pandemic influenza virus strains would likely take too long to prevent devastating global morbidity and mortality. If DNA vaccines can be distributed and delivered efficiently, genetic immunization could be an attractive solution to this problem, since plasmid DNA is stable, easily engineered to encode new protein antigens, and able to be quickly produced in large quantities. METHODS: We compared two novel genetic immunization methods in a mouse model of influenza to evaluate protective effects: aerosol delivery of polyethylenimine (PEI)-complexed hemagglutinin (HA)-expressing plasmid and intravenous (IV) delivery of the plasmid complexed with macroaggregated albumin/PEI. Serial serum samples were obtained for assay of neutralizing antibodies against HA. Mice were then challenged in the airway with influenza virus, and production of infectious virus in the lungs was titered. RESULTS: Most mice immunized with HA plasmid alone by aerosol and all mice immunized IV developed protective immune responses, whereas none administered control plasmid were protected. Aerosol co-administration of HA plasmid with plasmids encoding the cytokines interleukin 12 (IL12) and granulocyte-macrophage colony stimulating factor (GM-CSF) markedly increased neutralizing antibody responses, so that all aerosol immunized mice were protected from high level virus proliferation. CONCLUSIONS: Cytokine-enhanced aerosol delivery of plasmid vaccines can elicit robust protective immune responses against influenza. Thus, aerosol delivery has the potential to address the need for rapid widespread immunization against new influenza virus strains, and may have applications for other infectious and toxic disease processes.