Mapping of a protein-RNA kissing hairpin interface: Rom and Tar-Tar*.

An RNA 'kissing' complex is formed by the association of two hairpins via base pairing of their complementary loops. This sense-antisense RNA motif is used in the regulation of many cellular processes, including Escherichia coli ColE1 plasmid copy number. The RNA one modulator protein (Rom) acts as a co-regulator of ColE1 plasmid copy number by binding to the kissing hairpins and stabilizing their interaction. We have used heteronuclear two-dimensional NMR spectroscopy to map the interface between Rom and a kissing complex formed by the loop of the trans -activation response (Tar) element of immunodeficiency virus 1 (HIV-1) and its complement. The protein binding interface was obtained from changes in amide proton signals of uniformly 15N-labeled Rom with increasing concentrations of unlabeled Tar-Tar*. Similarly, the RNA-binding interface was obtained from changes in imino proton signals of uniformly 15N-labeled Tar with increasing concentrations of unlabeled Rom. Our results are in agreement with previous mutagenesis studies and provide additional information on Rom residues involved in RNA binding. The kissing hairpin interface with Rom leads to a model in which the protein contacts the minor groove of the loop-loop helix and, to a lesser extent, the major groove of the stems.     

Loss of phylogenetic information in chorion gene families of Bombyx mori gene conversion

The silkmoth chorion has provided a stimulating model for the study of evolution and developmental regulation of gene families. Previous attempts at inferring relationships among chorion sequences have been based on pairwise comparisons of overall similarity, a potentially problematic approach. To remedy this, we identified the alignable regions of low sequence variability and then analyzed this restricted database by parsimony and neighbor-joining methods. At the deepest level, the chorion sequence tree is split into two branches, called "alpha" and "beta." Within each branch, early- and late-expressing genes each constitute monophyletic groups, while the situation with middle-expressing genes remains uncertain. The HcB gene family appears to be the most basal beta-branch group, but this conclusion is qualified because the effect of gene conversion on branching order is unknown. Previous studies by Eickbush and colleagues have strongly suggested that ErA, HcA, and HcB families undergo gene conversion within a gene family, whereas the ErB family does not. The occurrence of conversion correlates with a particular tree structure; namely, branch lengths are much greater at the base of the family than at higher internodes and terminal branches. These observations raise the possibility that chorion gene families are defined by gene conversion events (reticulate evolution) rather than by descent with modification (synapomorphy).      

Identification of precursor to cytomegalovirus capsid assembly protein and evidence that processing results in loss of its carboxy-terminal end.

The 37-kilodalton (kDa) assembly protein of cytomegalovirus (strain Colburn) B capsids is shown to have a 40-kDa precursor. Pulse-chase radiolabeling experiments revealed that conversion of the precursor to the product was slow, requiring over 6 h for completion, and correlated with movement from the cytoplasmic to the nuclear fraction of Nonidet P-40-disrupted cells. Of these two proteins, only the 40-kDa precursor was synthesized in vitro from infected-cell RNA, consistent with its being the primary translation product. Amino acid sequence data obtained from CNBr-treated, high-performance liquid chromatography-purified assembly protein indicated that precursor translation begins at the first of two closely spaced potential initiation sites and that precursor maturation involves the loss of at least 32 amino acids from its carboxy-terminal end. It is also shown by immunological cross-reactivity and peptide similarity that three low-abundance B-capsid proteins (i.e., the 45-kilodalton [45K], 39K, and 38K proteins) are closely related to the assembly protein; the nature of this relatedness is discussed.     

First record of a spiral form of Aulacoseira,A. ambigua f. japonica (F.Meister) Tuji & D.M.Williams,in South African fresh waters

Water quality and algal populations in the Vaal River Barrage Reservoir have been monitored extensively for many decades, because of its importance as a water source for the most densely populated area in South Africa. Although Aulacoseira granulata (Ehrenberg) Simonsen is frequently found and at it times dominates algal assemblages in the Barrage, Aulacoseira ambigua (F.Meister) Tuji & D.M.Williams has never before been recorded at this locality. During a countrywide proficiency-testing scheme for algae counting, coordinated by Rand Water, spiral-shaped diatom colonies with distinctly curved cells were detected during May 2015. Upon investigation, it was found that the spiral colonies were Aulacoseira ambigua f. japonica, its presence in the fresh waters of South Africa being recorded for the first time. Since their first appearance during May 2015, colonies of Aulacoseira ambigua f. japonica have been constantly present in the surface waters of the Barrage. Their presence at this site can be linked to increasing eutrophication in the Vaal River, because the spiral form is known to prefer eutrophic conditions. Centric diatoms rarely form spiral colonies and there is some confusion in the literature as to the identity of this particular form, therefore in this paper we discuss this taxon, its nomenclature and ecological significance.     

Measurement with microelectrodes of intracellular Na+, K+, H+ and Cl- activities and of membrane potential in normal rat liver slices and during the 4-dimethylaminoazobenzene-induced rat hepatocarcinogenesis

Steady-state membrane potentials (Vm) and intracellular Na+ (aiNa), K+ (aiK), H+ (aiH) and Cl- (aiCl) activities were measured with double-barrelled ion-selective microelectrodes in liver slices from normal rats and during the 4-dimethylaminoazobenzene-induced (DAB) hepatocarcinogenesis. Rats fed with the experimental regimen without the carcinogen were used as control animals. In Krebs-Henseleit bicarbonate saline containing 5.5 mM glucose as bathing solution at 37 degrees C, Vm was found to be significantly lower in neoplastic hepatocytes, compared to normal liver cells. Vm decreased also in control rat liver cells. Increased Na+/K+ ratios and Na+ + K+ activities were found in cancerous hepatocytes whereas H+ and Cl- activities decreased. Therefore, the intracellular pH increased significantly in neoplastic cells, compared to normal and control cells. This could reflect activation of the Na+/H+ exchange system during the DAB-induced hepatocarcinogenesis, leading to a stimulation of cell metabolism with increased rate of protein and DNA synthesis and loss of growth control, under these conditions.     

Oligomerization states of the association domain and the holoenyzme of Ca2+/CaM kinase II

Ca2+/calmodulin activated protein kinase II (CaMKII) is an oligomeric protein kinase with a unique holoenyzme architecture. The subunits of CaMKII are bound together into the holoenzyme by the association domain, a C-terminal region of approximately 140 residues in the CaMKII polypeptide. Single particle analyses of electron micrographs have suggested previously that the holoenyzme forms a dodecamer that contains two stacked 6-fold symmetric rings. In contrast, a recent crystal structure of the isolated association domain of mouse CaMKIIalpha has revealed a tetradecameric assembly with two stacked 7-fold symmetric rings. In this study, we have determined the crystal structure of the Caenorhabditis elegans CaMKII association domain and it too forms a tetradecamer. We also show by electron microscopy that in its fully assembled form the CaMKII holoenzyme is a dodecamer but without the kinase domains, either from expression of the isolated association domain in bacteria or following their removal by proteolysis, the association domains form a tetradecamer. We speculate that the holoenzyme is held in its 6-fold symmetric state by the interactions of the N-terminal approximately 1-335 residues and that the removal of this region allows the association domain to convert into a more stable 7-fold symmetric form.     

7th SOSORT consensus paper: conservative treatment of idiopathic & Scheuermann's kyphosis

Abstract

Thoracic hyperkyphosis is a frequent problem and can impact greatly on patient's quality of life during adolescence. This condition can be idiopathic or secondary to Scheuermann disease, a disease disturbing vertebral growth. To date, there is no sound scientific data available on the management of this condition. Some studies discuss the effects of bracing, however no guidelines, protocols or indication's of treatment for this condition were found. The aim of this paper was to develop and verify the consensus on managing thoracic hyperkyphosis patients treated with braces and/or physiotherapy.

Methods

The Delphi process was utilised in four steps gradually modified according to the results of a set of recommendations: we involved the SOSORT Board twice, then all SOSORT members twice, with a Pre-Meeting Questionnaire (PMQ), and during a Consensus Session at the SOSORT Lyon Meeting with a Meeting Questionnaire (MQ).

Results

There was an unanimous agreement on the general efficacy of bracing and physiotherapy for this condition. Most experts suggested the use of 4-5 point bracing systems, however there was some controversy with regards to physiotherapeutic aims and modalities.

Conclusion

The SOSORT panel of experts suggest the use of rigid braces and physiotherapy to correct thoracic hyperkyphosis during adolescence. The evaluation of specific braces and physiotherapy techniques has been recommended.     

An optimized microarray platform for assaying genomic variation in <Emphasis Type="Italic">Plasmodium falciparum</Emphasis> field populations

We present an optimized probe design for copy number variation (CNV) and SNP genotyping in the Plasmodium falciparum genome. We demonstrate that variable length and isothermal probes are superior to static length probes. We show that sample preparation and hybridization conditions mitigate the effects of host DNA contamination in field samples. The microarray and workflow presented can be used to identify CNVs and SNPs with 95% accuracy in a single hybridization, in field samples containing up to 92% human DNA contamination.     

Removing high contrast artifacts via digital inpainting in cryo-electron tomography: an application of compressed sensing

To cope with poor quality in cryo-electron tomography images, electron-dense markers, such as colloidal goldbeads, are often used to assist image registration and analysis algorithms. However, these markers can create artifacts that occlude a specimen due to their high contrast, which can also cause failure of some image processing algorithms. One way of reducing these artifacts is to replace high contrast objects with pixel densities that blend into the surroundings in the projection domain before volume reconstruction. In this paper, we propose digital inpainting via compressed sensing (CS) as a new method to achieve this goal. We show that cryo-ET projections are sparse in the discrete cosine transform (DCT) domain, and, by finding the sparsest DCT domain decompositions given uncorrupted pixels, we can fill in the missing pixel values that are occluded by high contrast objects without discontinuities. Our method reduces visual artifacts both in projections and in tomograms better than conventional algorithms, such as polynomial interpolation and random noise inpainting.