Structure,diversity and evolutionary patterns of expressed MHC class II<Emphasis Type="Italic">B</Emphasis> genes in chub (<Emphasis Type="Italic">Squalius cephalus</Emphasis>), a cyprinid fish species from Europe

The polymorphism of exon 2 of the DAB genes (major histocompatibility complex [MHC] class IIB) was investigated for the first time in the freshwater cyprinid fish species, Squalius cephalus, in the wide range of its distribution in Europe. We identified 111 different MHC class IIB variants in 15 chub populations distributed from Finland to Spain. The sequence analysis showed that many structurally important amino acid sites that were conserved among tetrapods were also conserved in chub. The analysis of recombination indicated that it does not play an important role in producing and maintaining the variation of DAB genes analyzed in the present study. The exon 2 was shown to be subjected to intense positive selection. Phylogenetic analysis and sequence identities suggest the presence of two class IIB loci (DAB1-like and DAB3-like) in chub. Nevertheless, the presence of three DAB3-like sequence variants in several individuals indicates the duplication of the DAB3 gene. A contrasting selection pattern was found in DAB1-like and DAB3-like genes, which suggests the potential functional differences between these genes. Some DAB sequence variants were shared among the populations of different mtDNA lineages. The phylogenetic analyses did not confirm any biogeographical pattern of the genetic structure of MHC IIB in chub, which is in line with balancing selection and trans-species polymorphism in MHC genes. Nevertheless, cluster analysis based on the presence/absence of DAB sequence variants in the populations showed the phylogeophraphical pattern corresponding to the mtDNA lineages, which indicates that neutral selection can partially explain the MHC IIB evolution in chub.     

Life cycle of tortoise tick <Emphasis Type="Italic">Hyalomma aegyptium</Emphasis> under laboratory conditions

The tortoise tick Hyalomma aegyptium has a typical three-host life-cycle. Whereas its larvae and nymphs are less host-specific feeding on a variety of tetrapods, tortoises of the genus Testudo are principal hosts of adults. Ticks retained this trait also in our study under laboratory conditions, while adults were reluctant to feed on mammalian hosts. Combination of feeding larvae and nymphs on guinea pigs and feeding of adults on Testudo marginata tortoises provided the best results. Feeding period of females was on average 25 days (range 17–44), whereas males remain after female engorgement on tortoise host. Female pre-oviposition period was 14 days (3–31), followed by 24 days of oviposition (18–29). Pre-eclosion and eclosion, both together, takes 31 days (21–43). Larvae fed 5 days (3–9), then molted to nymphs after 17 days (12–23). Feeding period of nymphs lasted 7 days (5–10), engorged nymphs molted to adults after 24 days (19–26). Sex ratio of laboratory hatched H. aegyptium was nearly equal (1:1.09). The average weight of engorged female was 0.95 (0.72–1.12) g. The average number of laid eggs was 6,900 (6,524–7,532) per female, it was significantly correlated with weight of engorged female. Only 2.8% of engorged larvae and 1.8% of engorged nymphs remained un-molted and died. Despite the use of natural host species, feeding success of females reached only 45%. The whole life-cycle was completed within 147 days (98–215).     

The Negative Effect of Soy Extract on Erythrocyte Membrane Fluidity: An Electron Paramagnetic Resonance Study

A decrease of erythrocyte membrane fluidity can contribute to the pathophysiology of hypertension. Soy products, which are used as alternative therapeutics in some cardiovascular conditions, contain various isoflavones (genistein, daidzein, and their glucosides, genistin and daidzin), which can incorporate cellular membrane and change its fluidity. The aim of this study was to examine the effects of soy extract (which generally corresponds to the soy products of isoflavone composition) on erythrocyte membrane fluidity at graded depths. We used electron paramagnetic resonance spectroscopy and fatty acid spin probes (5-DS and 12-DS), the spectra of which are dependent on membrane fluidity. After being treated with soy extract, erythrocytes showed a significant (P = 0.016) decrease of membrane fluidity near the hydrophilic surface, while there were no significant changes of fluidity in deeper hydrophobic membrane regions. These results suggest that soy products containing high levels of genistein and isoflavone glucosides may not be suitable for use in hypertension because they decrease erythrocyte membrane fluidity.     

Molecular Variability and Evolution of the Pectate Lyase (<Emphasis Type="Italic">pel</Emphasis>-2) Parasitism Gene in Cyst Nematodes Parasitizing Different <Emphasis Type="Italic">Solanaceous</Emphasis> Plants

While pectate lyases are major parasitism factors in plant-parasitic nematodes, there is little information on the variability of these genes within species and their utility as pathotype or host range molecular markers. We have analysed polymorphisms of pectate lyase 2 (pel-2) gene, which degrades the unesterified polygalacturonate (pectate) of the host cell-wall, in the genus Globodera. Molecular variability of the pel-2 gene and the predicted protein was evaluated in populations of G. rostochiensis, G. pallida, G. “mexicana” and G. tabacum. Seventy eight pel-2 sequences were obtained and aligned. Point mutations were observed at 373 positions, 57% of these affect the coding part of the gene and produce 129 aa replacements. The observed polymorphism does not correlate either to the pathotypes proposed in potato cyst nematodes (PCN) or the subspecies described in tobacco cyst nematodes. The trees reveal a topology different from the admitted species topology as G. rostochiensis and G. pallida sequences are more similar to each other than to G. tabacum. Species-specific sites, potentially applicable for identification, and sites distinguishing PCN from tobacco cyst nematodes, were identified. As both G. rostochiensis and G. pallida display the same host range, but distinct from G. tabacum, which cannot parasitize potato plants, it is tempting to speculate that pel-2 genes polymorphism may be implicated in this adaptation, a view supported by the fact that no active pectate lyase 2 was found in G. “mexicana”, a close relative of G. pallida that is unable to develop on cultivated potato varieties.     

Human interleukin‐23 receptor antagonists derived from an albumin‐binding domain scaffold inhibit IL‐23‐dependent ex vivo expansion of IL‐17‐producing T‐cells

Engineered combinatorial libraries derived from small protein scaffolds represent a powerful tool for generating novel binders with high affinity, required specificity and designed inhibitory function. This work was aimed to generate a collection of recombinant binders of human interleukin‐23 receptor (IL‐23R), which is a key element of proinflammatory IL‐23‐mediated signaling. A library of variants derived from the three‐helix bundle scaffold of the albumin‐binding domain (ABD) of streptococcal protein G and ribosome display were used to select for high‐affinity binders of recombinant extracellular IL‐23R. A collection of 34 IL‐23R‐binding proteins (called REX binders), corresponding to 18 different sequence variants, was used to identify a group of ligands that inhibited binding of the recombinant p19 subunit of IL‐23, or the biologically active human IL‐23 cytokine, to the recombinant IL‐23R or soluble IL‐23R‐IgG chimera. The strongest competitors for IL‐23R binding in ELISA were confirmed to recognize human IL‐23R‐IgG in surface plasmon resonance experiments, estimating the binding affinity in the sub‐ to nanomolar range. We further demonstrated that several REX variants bind to human leukemic cell lines K‐562, THP‐1 and Jurkat, and this binding correlated with IL‐23R cell‐surface expression. The REX125, REX009 and REX128 variants competed with the p19 protein for binding to THP‐1 cells. Moreover, the presence of REX125, REX009 and REX115 variants significantly inhibited the IL‐23‐driven expansion of IL‐17‐producing primary human CD4⁺ T‐cells. Thus, we conclude that unique IL‐23R antagonists derived from the ABD scaffold were generated that might be useful in designing novel anti‐inflammatory biologicals. Proteins 2014; 82:975–989. © 2013 The Authors. Proteins: Structure, Function, and Bioinformatics Published by Wiley Periodicals, Inc.     

Proteomic Identification of a Candidate Sequence of Wheat Cytokinin-Binding Protein 1

Our knowledge on the principle mechanisms of cytokinin action has been significantly deepened over the last years, but several weakly explored areas still remain on the map of cytokinin cellular physiology. Cytokinin-binding proteins could also be included in this pending field of cytokinin research. Probably, the best explored representative of this group is the wheat cytokinin-binding protein 1 (CBP-1). The role of this germ-allocated protein as a presumable regulator of free aromatic cytokinin levels during grain germination has been discussed intensively. To dig deeper into this interesting protein, this study was aimed at the identification of the CBP-1 amino acid sequence. A combination of in silico BLAST search, classical biochemical CBP-1 purification based on isoelectric point precipitation and ion-exchange chromatography, and proteomic analysis of the isolated protein by ultra-high resolution tandem mass spectrometry allowed us to uncover and validate two CBP-1 subunit candidate sequences with molecular masses of 56.2 and 55.0 kDa, respectively. Interestingly, we found the latter sequence alternated in two amino acids in the putative cytokinin-binding motive in comparison to the composition of this domain reported in the original studies. A BLAST search for the amino acid sequence of the binding region among plant proteomes revealed several highly related protein sequences, all originating from the Poaceae family. This piece of information could give support to the elucidation of the role of CBP-1 in physiological processes mediated by aromatic cytokinins.     

Does a linear position transducer placed on a stick and belt provide sufficient validity and reliability of countermovement jump performance outcomes?

Manufacturers recommend that linear position transducers (LPTs) should be placed on the side of a barbell (or wooden dowel) to measure countermovement jump (CMJ) height, but the validity and reliability of this placement have not been compared to other attachment sites. Since this recommended attachment site is far from the centre of mass, a belt attachment where the LPT is placed between the feet may increase the validity and reliability of CMJ data. Thirty-six physical education students participated in the study (24.6 ± 4.3 years; 177.0 ± 7.7 cm; 77.2 ± 9.0 kg). Parameters from the two LPT attachments (barbell and belt) were simultaneously validated to force plate data, where the nature of bias was analysed (systematic vs random). The within-session and between-session reliability of both attachment sites were compared to force plate data using a test-retest protocol of two sets of 5 CMJs separated by 7 days. The LPT provided highly reliable and valid measures of peak force, mean force, mean power, and jump height, where the bias was mostly systematic (r² > 0.7; ICC > 0.9). Peak velocity, mean velocity, and peak power were in very good agreement with the force plate and were highly reliable (r² > 0.5; ICC > 0.7). Therefore, both attachment sites produced similar results with a systematic bias compared to force plate data. Thus, both attachment sites seem to be valid for assessing CMJs when the measuring tool and site remain consistent across measurements. However, if LPT data are to be compared to force plate data, recalculation equations should be used.     

Extensive loss of translational genes in the structurally dynamic mitochondrial genome of the angiosperm <Emphasis Type="Italic">Silene latifolia</Emphasis>

Background  

Mitochondrial gene loss and functional transfer to the nucleus is an ongoing process in many lineages of plants, resulting in substantial variation across species in mitochondrial gene content. The Caryophyllaceae represents one lineage that has experienced a particularly high rate of mitochondrial gene loss relative to other angiosperms.