Segment TM7 from the cytoplasmic hemi-channel from VO-H+-V-ATPase includes a flexible region that has a potential role in proton translocation

A 900-MHz NMR study is reported of peptide sMTM7 that mimics the cytoplasmic proton hemi-channel domain of the seventh transmembrane segment (TM7) from subunit a of H(+)-V-ATPase from Saccharomyces cerevisiae. The peptide encompasses the amino acid residues known to actively participate in proton translocation. In addition, peptide sMTM7 contains the amino acid residues that upon mutation cause V-ATPase to become resistant against the inhibitor bafilomycin. 2D TOCSY and NOESY (1)H-(1)H NMR spectra are obtained of sMTM7 dissolved in d(6)-DMSO and are used to calculate the three-dimensional structure of the peptide. The NMR-based structures and corresponding dynamical features of peptide sMTM7 show that sMTM7 is composed of two alpha-helical regions. These regions are separated by a flexible hinge of two residues. The hinge acts as a ball-and-joint socket and both helical segments move independently with respect to one another. This movement in TM7 is suggested to cause the opening and closing of the cytoplasmic proton hemi-channel and enables proton translocation.     

Phylogenetic screening of the human genome: identification of differentially hybridizing repetitive sequence families

The phi-screen, a method of phylogenetic screening, can be employed to detect repetitive sequence families that differentially hybridize between closely related species. Such differences may involve sequence divergence or variations in copy number, including total presence versus absence of a family of repeated DNA. We present the results of a phi-screen comparing the human genome to that of the prosimian, Galago crassicaudatus. Three human repetitive families that are divergent or not present in galago have been detected. One of these families is described in detail; it is similar among the anthropoids but is present in a lower copy number and/or divergent form in prosimians. The family is clearly related to the transposon-like human element (THE) described by Paulson et al. (1985). THEs have long terminal repeats reminiscent of retroviruses but are unique in that they have no sequence similarity to known mammalian retroviruses. The sequence of a solo long terminal repeat, found unassociated with THE internal sequence, is presented. This family member, THE p2, is bordered by a 5-bp target-site repeat and is interrupted by the insertion of an Alu element. A solo THE element sequenced by Wiginton et al. (1986) contains an insertion of Alu at precisely the same position as does THE p2.      

RET/GFRα Signals Are Dispensable for Thymic T Cell Development In Vivo

Identification of thymocyte regulators is a central issue in T cell biology. Interestingly, growing evidence indicates that common key molecules control neuronal and immune cell functions. The neurotrophic factor receptor RET mediates critical functions in foetal hematopoietic subsets, thus raising the possibility that RET-related molecules may also control T cell development. We show that Ret, Gfra1 and Gfra2 are abundantly expressed by foetal and adult immature DN thymocytes. Despite the developmentally regulated expression of these genes, analysis of foetal thymi from Gfra1, Gfra2 or Ret deficient embryos revealed that these molecules are dispensable for foetal T cell development. Furthermore, analysis of RET gain of function and Ret conditional knockout mice showed that RET is also unnecessary for adult thymopoiesis. Finally, competitive thymic reconstitution assays indicated that Ret deficient thymocytes maintained their differentiation fitness even in stringent developmental conditions. Thus, our data demonstrate that RET/GFRα signals are dispensable for thymic T cell development in vivo, indicating that pharmacological targeting of RET signalling in tumours is not likely to result in T cell production failure.     

The Role of Efflux Pumps in Schistosoma mansoni Praziquantel Resistant Phenotype

Background

Schistosomiasis is a neglected disease caused by a trematode of the genus Schistosoma that is second only to malaria in public health significance in Africa, South America, and Asia. Praziquantel (PZQ) is the drug of choice to treat this disease due to its high cure rates and no significant side effects. However, in the last years increasingly cases of tolerance to PZQ have been reported, which has caused growing concerns regarding the emergency of resistance to this drug.

Methodology/Principal Findings

Here we describe the selection of a parasitic strain that has a stable resistance phenotype to PZQ. It has been reported that drug resistance in helminths might involve efflux pumps such as members of ATP-binding cassette transport proteins, including P-glycoprotein and multidrug resistance-associated protein families. Here we evaluate the role of efflux pumps in Schistosoma mansoni resistance to PZQ, by comparing the efflux pumps activity in susceptible and resistant strains. The evaluation of the efflux activity was performed by an ethidium bromide accumulation assay in presence and absence of Verapamil. The role of efflux pumps in resistance to PZQ was further investigated comparing the response of susceptible and resistant parasites in the absence and presence of different doses of Verapamil, in an ex vivo assay, and these results were further reinforced through the comparison of the expression levels of SmMDR2 RNA by RT-PCR.

Conclusions/Significance

This work strongly suggests the involvement of Pgp-like transporters SMDR2 in Praziquantel drug resistance in S. mansoni. Low doses of Verapamil successfully reverted drug resistance. Our results might give an indication that a combination therapy with PZQ and natural or synthetic Pgp modulators can be an effective strategy for the treatment of confirmed cases of resistance to PZQ in S. mansoni.     

Numerical response of predators to large variations of grassland vole abundance and long‐term community changes

Voles can reach high densities with multiannual population fluctuations of large amplitude, and they are at the base of predator communities in Northern Eurasia and Northern America. This status places them at the heart of management conflicts wherein crop protection and health concerns are often raised against conservation issues. Here, a 20‐year survey describes the effects of large variations in grassland vole populations on the densities and the daily theoretical food intakes (TFI) of vole predators based on roadside counts. Our results show how the predator community responded to prey variations of large amplitude and how it reorganized with the increase in a dominant predator, here the red fox, which likely negatively impacted hare, European wildcat, and domestic cat populations. This population increase did not lead to an increase in the average number of predators present in the study area, suggesting compensations among resident species due to intraguild predation or competition. Large variations in vole predator number could be clearly attributed to the temporary increase in the populations of mobile birds of prey in response to grassland vole outbreaks. Our study provides empirical support for more timely and better focused actions in wildlife management and vole population control, and it supports an evidence‐based and constructive dialogue about management targets and options between all stakeholders of such socio‐ecosystems.

A 20‐year survey describes the effects of large variations in grassland vole populations on the densities and the daily theoretical food intakes of a vole predator community based on roadside counts. Our results show how the predator community responds to prey variations of large amplitude and how it reorganized with the increase of the red fox, which likely negatively impacted hare, European wildcat, and domestic cat populations.     

Complete genome sequencing of a novel newcastle disease virus isolate circulating in layer chickens in the dominican republic

Newcastle disease virus (NDV) was isolated from an outbreak in layer chickens in the Dominican Republic in 2008. Infections with this isolate led to a 100% apparent case fatality rate in birds. Complete genome sequencing revealed that the isolate does not belong to any of the previously described NDV genotypes. Similarly, large differences were observed in the amino acid sequence of the fusion and hemagglutinin-neuraminidase proteins in comparison with all known NDV genotypes, suggesting the existence of an unknown reservoir for NDV. The work presented here represents the first complete genome sequence of NDV in the Dominican Republic.     

Creatine supplementation reduces oxidative stress biomarkers after acute exercise in rats

The objective of this study was to evaluate the effect of creatine supplementation on muscle and plasma markers of oxidative stress after acute aerobic exercise. A total of 64 Wistar rats were divided into two groups: control group (n = 32) and creatine-supplemented group (n = 32). Creatine supplementation consisted of the addition of 2% creatine monohydrate to the diet. After 28 days, the rats performed an acute moderate aerobic exercise bout (1-h swimming with 4% of total body weight load). The animals were killed before (pre) and at 0, 2 and 6 h (n = 8) after acute exercise. As expected, plasma and total muscle creatine concentrations were significantly higher (P < 0.05) in the creatine-supplemented group compared to control. Acute exercise increased plasma thiobarbituric acid reactive species (TBARS) and total lipid hydroperoxide. The same was observed in the soleus and gastrocnemius muscles. Creatine supplementation decreased these markers in plasma (TBARS: pre 6%, 0 h 25%, 2 h 27% and 6 h 20%; plasma total lipid hydroperoxide: pre 38%, 0 h 24%, 2 h 12% and 6 h 20%, % decrease). Also, acute exercise decreased the GSH/GSSG ratio in soleus muscle, which was prevented by creatine supplementation (soleus: pre 8%, 0 h 29%, 2 h 30% and 6 h 44%, % prevention). The results show that creatine supplementation inhibits increased oxidative stress markers in plasma and muscle induced by acute exercise.     

Hormones that increase maternal responsiveness affect accumbal dopaminergic responses to pup- and food-stimuli in the female rat

The present study investigated hormonal mediation of maternal behavior and accumbal dopamine (DA) responses to pup-stimuli, as measured in microdialysis samples collected from the nucleus accumbens shell of female rats in non-homecage environment. In Experiment 1, samples were collected before and after continuous homecage pup experience from either intact postpartum or cycling females. In Experiment 2, samples were collected before and after responding maternally in homecage from ovariectomized females given either parturient-like hormone or sham treatments. After baseline sample collection in the dialysis chamber, pup and food stimuli were individually presented to females. Upon sampling completion, all animals were placed back into their homecage with donor pups for several days, and then the sample collection procedure was repeated. Prior to stimulus presentation, postpartum and hormone-treated females had decreased basal DA release compared to their controls. In response to pup stimuli, only postpartum and hormone-treated females had increased DA release compared to basal release (both sampling days). In response to food stimuli, all females had increased DA responses from basal; although there were group differences on the initial day of sampling. Findings suggest that hormones associated with inducing maternal behavior in the postpartum rat play a significant role in modifying accumbal dopaminergic responses on first exposure to pup stimuli in the rat. However, the postpartum experience provides further modifications to this brain region to promote DA responses to pup stimuli.     

Genetic diversity and geographic differentiation of disjunct Atlantic and Amazonian populations of <Emphasis Type="Italic">Psychotria ipecacuanha</Emphasis> (Rubiaceae)

Ipecac (Psychotria ipecacuanha) is a perennial, medicinal herb that grows in the understory of semi-deciduous tropical forests in the Neotropics. Ipecacs present a widely disjunct distribution, with two of its three ranges occurring in Brazil. The Amazonian populations are at least 1600 km from the nearest Atlantic populations. This work used ISSR markers to compare the genetic diversity and structure of populations from the two Brazilian ranges. Lower genetic diversity in Amazon populations (P = 60.11%, Hs = 0.18) and higher genetic diversity in Atlantic populations (P = 73.94%, Hs = 0.20) were detected. Differentiation between ranges were high (θ ^B = 0.6838, G_ST-B = 0.6665). AMOVA revealed that 65.3% of the total molecular variance can be attributed to regional differences between the two ranges. Principal coordinate analyses and cluster analyses organized ipecacs at either individual or population level into two exclusive groups that correspond each to one of the two disjunct ranges, without exception. The results do not support a scenario that postulates human-mediated, long-distance dispersal events as a plausible origin for the distribution of the Brazilian ipecacs, but indicate geographic isolation as a long-standing barrier to genetic exchange and connectivity among populations from different ranges. Conservation implications are discussed.     

NRP/Optineurin Cooperates with TAX1BP1 to Potentiate the Activation of NF-κB by Human T-Lymphotropic Virus Type 1 Tax Protein

Nuclear factor (NF)-κB is a major survival pathway engaged by the Human T-Lymphotropic Virus type 1 (HTLV-1) Tax protein. Tax1 activation of NF-κB occurs predominantly in the cytoplasm, where Tax1 binds NF-κB Essential Modulator (NEMO/IKKγ) and triggers the activation of IκB kinases. Several independent studies have shown that Tax1-mediated NF-κB activation is dependent on Tax1 ubiquitination. Here, we identify by co-immunoprecipitation assays NEMO-Related Protein (NRP/Optineurin) as a binding partner for Tax1 in HTLV-1 infected and Tax1/NRP co-expressing cells. Immunofluorescence studies reveal that Tax1, NRP and NEMO colocalize in Golgi-associated structures. The interaction between Tax1 and NRP requires the ubiquitin-binding activity of NRP and the ubiquitination sites of Tax1. In addition, we observe that NRP increases the ubiquitination of Tax1 along with Tax1-dependent NF-κB signaling. Surprisingly, we find that in addition to Tax1, NRP interacts cooperatively with the Tax1 binding protein TAX1BP1, and that NRP and TAX1BP1 cooperate to modulate Tax1 ubiquitination and NF-κB activation. Our data strongly suggest for the first time that NRP is a critical adaptor that regulates the assembly of TAX1BP1 and post-translationally modified forms of Tax1, leading to sustained NF-κB activation.