snoRNPs Regulate Telomerase Activity in Neuroblastoma and Are Associated with Poor Prognosis

Amplification of the MYCN oncogene is strongly associated with poor prognosis in neuroblastoma (NB). In addition to MYCN amplification, many studies have focused on identifying patients with a poor prognosis based on gene expression profiling. The majority of prognostic signatures today are comprised of large gene lists limiting their clinical application. In addition, although of prognostic significance,most of these signatures fail to identify cellular processes that can explain their relation to prognosis. Here, we determined prognostically predictive genes in a data set containing 251 NBs. Gene Ontology analysis was performed on significant genes with a positive hazard ratio to search for cellular processes associated with poor prognosis. An enrichment in ribonucleoproteins (RNPs) was found. Genes involved in the stabilization and formation of the central small nucleolar RNP (snoRNP) complex were scrutinized using a backward conditional Cox regression resulting in an snoRNP signature consisting of three genes: DKC1, NHP2, and GAR1. The snoRNP signature significantly and independently predicted prognosis when compared to the established clinical risk factors. Association of snoRNP protein expression and prognosis was confirmed using tissue microarrays. Knockdown of snoRNP expression in NB cell lines resulted in reduced telomerase activity and an increase in anaphase bridge frequency. In addition, in patient material, expression of the snoRNP complex was significantly associated with telomerase activity, occurrence of segmental aberrations, and expression-based measurements of chromosomal instability. Together, these results underscore the prognostic value of snoRNP complex expression in NB and suggest a role for snoRNPs in telomere maintenance and genomic stability.     

Inhibition of the Smc5/6 Complex during Meiosis Perturbs Joint Molecule Formation and Resolution without Significantly Changing Crossover or Non-crossover Levels

Meiosis is a specialized cell division used by diploid organisms to form haploid gametes for sexual reproduction. Central to this reductive division is repair of endogenous DNA double-strand breaks (DSBs) induced by the meiosis-specific enzyme Spo11. These DSBs are repaired in a process called homologous recombination using the sister chromatid or the homologous chromosome as a repair template, with the homolog being the preferred substrate during meiosis. Specific products of inter-homolog recombination, called crossovers, are essential for proper homolog segregation at the first meiotic nuclear division in budding yeast and mice. This study identifies an essential role for the conserved Structural Maintenance of Chromosomes (SMC) 5/6 protein complex during meiotic recombination in budding yeast. Meiosis-specific smc5/6 mutants experience a block in DNA segregation without hindering meiotic progression. Establishment and removal of meiotic sister chromatid cohesin are independent of functional Smc6 protein. smc6 mutants also have normal levels of DSB formation and repair. Eliminating DSBs rescues the segregation block in smc5/6 mutants, suggesting that the complex has a function during meiotic recombination. Accordingly, smc6 mutants accumulate high levels of recombination intermediates in the form of joint molecules. Many of these joint molecules are formed between sister chromatids, which is not normally observed in wild-type cells. The normal formation of crossovers in smc6 mutants supports the notion that mainly inter-sister joint molecule resolution is impaired. In addition, return-to-function studies indicate that the Smc5/6 complex performs its most important functions during joint molecule resolution without influencing crossover formation. These results suggest that the Smc5/6 complex aids primarily in the resolution of joint molecules formed outside of canonical inter-homolog pathways.     

A Large Plasmodium vivax Reservoir and Little Population Structure in the South Pacific

Introduction

The importance of Plasmodium vivax in malaria elimination is increasingly being recognized, yet little is known about its population size and population genetic structure in the South Pacific, an area that is the focus of intensified malaria control.

Methods

We have genotyped 13 microsatellite markers in 295 P. vivax isolates from four geographically distinct sites in Papua New Guinea (PNG) and one site from Solomon Islands, representing different transmission intensities.

Results

Diversity was very high with expected heterozygosity values ranging from 0.62 to 0.98 for the different markers. Effective population size was high (12′872 to 19′533 per site). In PNG population structuring was limited with moderate levels of genetic differentiation. F _ST values (adjusted for high diversity of markers) were 0.14–0.15. Slightly higher levels were observed between PNG populations and Solomon Islands (F _ST = 0.16).

Conclusions

Low levels of population structure despite geographical barriers to transmission are in sharp contrast to results from regions of low P. vivax endemicity. Prior to intensification of malaria control programs in the study area, parasite diversity and effective population size remained high.     

Long-term temporal variation in the organization of an ant–plant network

Background and Aims

Functional groups of species interact and coevolve in space and time, forming complex networks of interacting species. A long-term study of temporal variation of an ant–plant network is presented with the aims of: (1) depicting its structural changes over a 20-year period; (2) detailing temporal variation in network topology, as revealed by nestedness and modularity analysis and other parameters (i.e. connectance, niche overlap); and (3) identifying long-term turnover in taxonomic structure (i.e. switches in ant resource use or plant visitor assemblages according to taxa).

Methods

Fieldwork was carried out at La Mancha, Mexico, and ant–plant interactions were observed between 1989 and 1991, between 1998 and 2000, and between May 2010 and 2011. Occurrences of ants on extrafloral nectaries (EFNs) were recorded. The resulting ant–plant networks were constructed from qualitative presence–absence data determined by a species–species matrix defined by the frequency of occurrence of each pairwise ant–plant interaction.

Key Results

Network variation across time was stable and a persistent nested structure may have contributed to the maintenance of resilient and species-rich communities. Modularity was lower than expected, especially in the most recent networks, indicating that the community exhibited high overlap among interacting species (e.g. few species were hubs in the more recent network, being partly responsible for the nested pattern). Structurally, the connections created among modules by super-generalists gave cohesion to subsets of species that otherwise would remain unconnected. This may have allowed an increasing cascade-effect of evolutionary events among modules. Mutualistic ant–plant interactions were structured 20 years ago mainly by the subdominant nectarivorous ant species Camponotus planatus and Crematogaster brevispinosa, which monopolized the best extrafloral nectar resources and out-competed other species with broader feeding habits. Through time, these ants, which are still present, lost their position as network hubs and diminished in their importance in structuring the network; simultaneously, plants gained in importance.

Conclusions

The long-term network analysis reveals a decrease in attended plant species richness, a notable increase in plant species participation from 1990 to 2010 (sustained by less plant taxonomic similarity in the older 1990 network), an increase in the number of ant species and a diminishing dominance of super-generalist ants. The structure of the community has remained highly nested and connected with low modularity, suggesting overall a more participative, homogeneous, cohesive interaction network. Although previous studies have suggested that interactions between ants and EFN-bearing plants are susceptible to seasonality, abiotic factors and perturbation, this cohesive structure appears to be the key for biodiversity and community maintenance.     

Esterase 2 as a fluorescent biosensor for the detection of organophosphorus compounds: docking and electronic insights from molecular dynamics

ABSTRACT

Organophosphorus compounds (OP) are mainly used in agriculture as pesticides. Unfortunately, each year many rural workers are intoxicated by these compounds and, many times, the diagnosis of the exact molecule causing the intoxication can be tardy, exposing the patients to a huge risk of death. One way of preventing this delay is the use of enzymatic biosensors like the enzyme Esterase 2 from Alicyclobacillus acidocaldarius (AaEST2), which is an efficient fluorescent biosensor for OP identification. However, although this enzyme has been well studied experimentally, the complete understanding of the energy transfer processes that occur between AaEST2 and OPs is still obscure, making it difficult the accurate identification of the OP. In order to better understand this process, we applied in this work molecular docking and molecular dynamics studies, together with the Förster fluorescence resonance energy transfer (FRET) theory, to achieve a better understanding of the fluorescence profiles that are described in the literature and correlate them to individual OPs. Our results suggest that the pesticides chlorpyrifos, diazinon, parathion and paraoxon are all capable of quenching the residue Trp85 from AaEST2, triggering fluorescence. This supports our hypothesis that AaEST2 can be used as a fluorescent biosensor for the detection of organophosphorus compounds.     

Minor contribution of leaf litter to N nutrition of beech (Fagus sylvatica) seedlings in a mountainous beech forest of Southern Germany

Aims

Our aims were to characterize the fate of leaf-litter-derived nitrogen in the plant-soil-microbe system of a temperate beech forest of Southern Germany and to identify its importance for N nutrition of beech seedlings.

Methods

¹⁵N-labelled leaf litter was traced in situ into abiotic and biotic N pools in mineral soil as well as into beech seedlings and mycorrhizal root tips over three growing seasons.

Results

There was a rapid transfer of ¹⁵N into the mineral soil already 21 days after tracer application with soil microbial biomass initially representing the dominant litter-N sink. However, ¹⁵N recovery in non-extractable soil N pools strongly increased over time and subsequently became the dominant ¹⁵N sink. Recovery in plant biomass accounted for only 0.025 % of ¹⁵N excess after 876 days. After three growing seasons, ¹⁵N excess recovery was characterized by the following sequence: non-extractable soil N?>>?extractable soil N including microbial biomass?>>?plant biomass?>?ectomycorrhizal root tips.

Conclusions

After quick vertical dislocation and cycling through microbial N pools, there was a rapid stabilization of leaf-litter-derived N in non-extractable N pools of the mineral soil. Very low ¹⁵N recovery in beech seedlings suggests a high importance of other N sources such as root litter for N nutrition of beech understorey.     

Characterization of a Serine Hydrolase Targeted by Acyl-protein Thioesterase Inhibitors in Toxoplasma gondii

In eukaryotic organisms, cysteine palmitoylation is an important reversible modification that impacts protein targeting, folding, stability, and interactions with partners. Evidence suggests that protein palmitoylation contributes to key biological processes in Apicomplexa with the recent palmitome of the malaria parasite Plasmodium falciparum reporting over 400 substrates that are modified with palmitate by a broad range of protein S-acyl transferases. Dynamic palmitoylation cycles require the action of an acyl-protein thioesterase (APT) that cleaves palmitate from substrates and conveys reversibility to this posttranslational modification. In this work, we identified candidates for APT activity in Toxoplasma gondii. Treatment of parasites with low micromolar concentrations of β-lactone- or triazole urea-based inhibitors that target human APT1 showed varied detrimental effects at multiple steps of the parasite lytic cycle. The use of an activity-based probe in combination with these inhibitors revealed the existence of several serine hydrolases that are targeted by APT1 inhibitors. The active serine hydrolase, TgASH1, identified as the homologue closest to human APT1 and APT2, was characterized further. Biochemical analysis of TgASH1 indicated that this enzyme cleaves substrates with a specificity similar to APTs, and homology modeling points toward an APT-like enzyme. TgASH1 is dispensable for parasite survival, which indicates that the severe effects observed with the β-lactone inhibitors are caused by the inhibition of non-TgASH1 targets. Other ASH candidates for APT activity were functionally characterized, and one of them was found to be resistant to gene disruption due to the potential essential nature of the protein.