Marker-free image registration of electron tomography tilt-series

Background  

Tilt series are commonly used in electron tomography as a means of collecting three-dimensional information from two-dimensional projections. A common problem encountered is the projection alignment prior to 3D reconstruction. Current alignment techniques usually employ gold particles or image derived markers to correctly align the images. When these markers are not present, correlation between adjacent views is used to align them. However, sequential pairwise correlation is prone to bias and the resulting alignment is not always optimal.     

Entomotoxic action of Sambucus nigra agglutinin i in Acyrthosiphon pisum aphids and Spodoptera exigua caterpillars through caspase‐3‐like‐dependent apoptosis

In this project, the toxicity and mechanism of action of the ricin‐B‐related lectin SNA‐I from elderberry (Sambucus nigra) in the pea aphid (Acyrthosiphon pisum) and the beet armyworm (Spodoptera exigua), two important pest insects in agriculture, were studied. SNA‐I is a chimeric lectin belonging to the class of ribosome‐inactivating proteins and consists of an A‐chain with N‐glycosidase activity and a carbohydrate‐binding B‐chain. Incorporation of 2 mg/ml of SNA‐I in the diet of neonates and adults of A. pisum caused 40–46% mortality within 2 days, while in third instars of S. exigua, the larval biomass was significantly reduced by 12% after feeding for 3 days on a diet containing 5 mg/g of SNA‐I. Interestingly, extracts of the (mid)gut of treated A. pisum and S. exigua demonstrated DNA fragmentation and this was accompanied with an increase in caspase‐3‐like activity. The involvement of cell death or apoptosis in the entomotoxicity of SNA‐I through induction of caspase‐3‐like activity was also confirmed by addition of the permeable caspase‐3 inhibitor III in the diet, leading to a rescue of the treated aphid neonates. Finally, similar to the chimeric lectin SNA‐I, the hololectin SNA‐II, consisting of two carbohydrate‐binding B‐chains caused high mortality to neonate A. pisum aphids with an LC₅₀ of 1.59 mg/ml, suggesting that the entomotoxic action of the lectins under study mainly relies on their carbohydrate‐binding activity. © 2010 Wiley Periodicals, Inc.     

The genomic landscape of the verrucomicrobial methanotroph Methylacidiphilum fumariolicum SolV

Background

Aerobic methanotrophs can grow in hostile volcanic environments and use methane as their sole source of energy. The discovery of three verrucomicrobial Methylacidiphilum strains has revealed diverse metabolic pathways used by these methanotrophs, including mechanisms through which methane is oxidized. The basis of a complete understanding of these processes and of how these bacteria evolved and are able to thrive in such extreme environments partially resides in the complete characterization of their genome and its architecture.

Results

In this study, we present the complete genome sequence of Methylacidiphilum fumariolicum SolV, obtained using Pacific Biosciences single-molecule real-time (SMRT) sequencing technology. The genome assembles to a single 2.5 Mbp chromosome with an average GC content of 41.5%. The genome contains 2,741 annotated genes and 314 functional subsystems including all key metabolic pathways that are associated with Methylacidiphilum strains, including the CBB pathway for CO₂ fixation. However, it does not encode the serine cycle and ribulose monophosphate pathways for carbon fixation. Phylogenetic analysis of the particulate methane mono-oxygenase operon separates the Methylacidiphilum strains from other verrucomicrobial methanotrophs. RNA-Seq analysis of cell cultures growing in three different conditions revealed the deregulation of two out of three pmoCAB operons. In addition, genes involved in nitrogen fixation were upregulated in cell cultures growing in nitrogen fixing conditions, indicating the presence of active nitrogenase. Characterization of the global methylation state of M. fumariolicum SolV revealed methylation of adenines and cytosines mainly in the coding regions of the genome. Methylation of adenines was predominantly associated with 5′-^m6ACN₄GT-3′ and 5′-CC^m6AN₅CTC-3′ methyltransferase recognition motifs whereas methylated cytosines were not associated with any specific motif.

Conclusions

Our findings provide novel insights into the global methylation state of verrucomicrobial methanotroph M. fumariolicum SolV. However, partial conservation of methyltransferases between M. fumariolicum SolV and M. infernorum V4 indicates potential differences in the global methylation state of Methylacidiphilum strains. Unravelling the M. fumariolicum SolV genome and its epigenetic regulation allow for robust characterization of biological processes that are involved in oxidizing methane. In turn, they offer a better understanding of the evolution, the underlying physiological and ecological properties of SolV and other Methylacidiphilum strains.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-914) contains supplementary material, which is available to authorized users.     

Lack of enantiomeric specificity in the effects of anesthetic steroids on lipid bilayers

The most important target protein for many anesthetics, including volatile and steroid anesthetics, appears to be the type A γ-amino butyric acid receptor (GABA_AR), yet direct binding remains to be demonstrated. Hypotheses of lipid-mediated anesthesia suggest that lipid bilayer properties are changed by anesthetics and that this in turn affects the functions of proteins. While other data could equally well support direct or lipid-mediated action, enantiomeric specificity displayed by some anesthetics is not reflected in their interactions with lipids. In the present study, we studied the effects of two pairs of anesthetic steroid enantiomers on bilayers of several compositions, measuring potentially relevant physical properties. For one of the pairs, allopregnanolone and ent-allopregnanolone, the natural enantiomer is 300% more efficacious as an anesthetic, while for the other, pregnanolone and ent-pregnanolone, there is little difference in anesthetic potency. For each enantiomer pair, we could find no differences. This strongly favors the view that the effects of these anesthetics on lipid bilayers are not relevant for the main features of anesthesia. These steroids also provide tools to distinguish in general the direct binding of steroids to proteins from lipid-mediated effects.