Cholesterol sensing by CD81 is important for hepatitis C virus entry

CD81 plays a central role in a variety of physiological and pathological processes. Recent structural analysis of CD81 indicates that it contains an intramembrane cholesterol-binding pocket and that interaction with cholesterol may regulate a conformational switch in the large extracellular domain of CD81. Therefore, CD81 possesses a potential cholesterol-sensing mechanism; however, its relevance for protein function is thus far unknown. In this study we investigate CD81 cholesterol sensing in the context of its activity as a receptor for hepatitis C virus (HCV). Structure-led mutagenesis of the cholesterol-binding pocket reduced CD81–cholesterol association but had disparate effects on HCV entry, both reducing and enhancing CD81 receptor activity. We reasoned that this could be explained by alterations in the consequences of cholesterol binding. To investigate this further we performed molecular dynamic simulations of CD81 with and without cholesterol; this identified a potential allosteric mechanism by which cholesterol binding regulates the conformation of CD81. To test this, we designed further mutations to force CD81 into either the open (cholesterol-unbound) or closed (cholesterol-bound) conformation. The open mutant of CD81 exhibited reduced HCV receptor activity, whereas the closed mutant enhanced activity. These data are consistent with cholesterol sensing switching CD81 between a receptor active and inactive state. CD81 interactome analysis also suggests that conformational switching may modulate the assembly of CD81–partner protein networks. This work furthers our understanding of the molecular mechanism of CD81 cholesterol sensing, how this relates to HCV entry, and CD81''s function as a molecular scaffold; these insights are relevant to CD81''s varied roles in both health and disease.     

Multi‐generational long‐distance migration of insects: studying the painted lady butterfly in the Western Palaearctic

Long‐range, seasonal migration is a widespread phenomenon among insects, allowing them to track and exploit abundant but ephemeral resources over vast geographical areas. However, the basic patterns of how species shift across multiple locations and seasons are unknown in most cases, even though migrant species comprise an important component of the temperate‐zone biota. The painted lady butterfly Vanessa cardui is such an example; a cosmopolitan continuously‐brooded species which migrates each year between Africa and Europe, sometimes in enormous numbers. The migration of 2009 was one of the most impressive recorded, and thousands of observations were collected through citizen science programmes and systematic entomological surveys, such as high altitude insect‐monitoring radar and ground‐based butterfly monitoring schemes. Here we use V. cardui as a model species to better understand insect migration in the Western Palaearctic, and we capitalise on the complementary data sources available for this iconic butterfly. The migratory cycle in this species involves six generations, encompassing a latitudinal shift of thousands of kilometres (up to 60 degrees of latitude). The cycle comprises an annual poleward advance of the populations in spring followed by an equatorward return movement in autumn, with returning individuals potentially flying thousands of kilometres. We show that many long‐distance migrants take advantage of favourable winds, moving downwind at high elevation (from some tens of metres from the ground to altitudes over 1000 m), pointing at strong similarities in the flight strategies used by V. cardui and other migrant Lepidoptera. Our results reveal the highly successful strategy that has evolved in these insects, and provide a useful framework for a better understanding of long‐distance seasonal migration in the temperate regions worldwide.     

Predation on snakes of Argentina: Effects of coloration and ring pattern on coral and false coral snakes

The occurrence of coral snake coloration among unrelated venomous and non‐venomous snake species has often been explained in terms of warning coloration and mimicry. In Argentina, no field tests have been conducted to confirm this mimetic association between one venomous coral species (Micrurus phyrrocryptus, Elapidae) and two non‐venomous snake species with a similar color pattern (Lystrophis pulcher and Oxyrhopus rhombifer, Colubridae). The aims of this work were to test for the possible aposematic or cryptic function of the ring pattern and coloration of coral snakes and false coral snakes from central Argentina, and to analyse whether the pattern is effective throughout the year. Predation on snakes was estimated by using non‐toxic plasticine replicas of ringed venomous and non‐venomous snakes and unbanded green snakes placed along transects in their natural habitat during the dry and rainy season. Ringed color pattern was attacked by predators despite the background color. One of the replica types was attacked more than expected during the dry season, suggesting that both shape and width of rings may influence the choice by predators. The reaction of predators towards replicas that mimic snake species with ringed patterns is independent of the geographical region, and we can conclude that mimicry characteristics are quite general when the true models are present in the area.     

Effects of Phytic Acid and Exercise on Some Serum Analytes in Rats Orally Exposed to Diets Supplemented with Cadmium

Cadmium is an environmental pollutant of increasing worldwide concern. It has been reported to be high in the soil where food crops are grown in some parishes of Jamaica. Surprisingly, no adverse effect of cadmium has been reported among the Jamaican population. However, phytic acid has also been shown to be high in some food crops grown in Jamaica. In this study, we evaluated the effects of phytic acid (1 %) and exercise on the metabolism of cadmium (5 mg cadmium/kg body weight) in rats. Five groups of rats were fed as follows: rats fed control diet, control diet supplemented with cadmium and subjected to exercise, control diet supplemented with phytic acid plus cadmium and subjected to exercise, control diet supplemented with cadmium plus phytic acid, and control diet supplemented with cadmium only. The animals were fed for 4 weeks and then sacrificed. Blood samples were collected for some biochemical assays. Percentage weight loss (28.42 %) was greatest in the group that had cadmium supplement only. The group fed control diet supplemented with cadmium only displayed increased liver enzymes and electrolytes except for the significant decrease in bicarbonate compared to other test groups. Similarly, blood urea nitrogen and uric acid were increased in the group fed cadmium supplement only compared to other test groups. Total cholesterol trended downwards in the test groups compared to control. These observations suggest that consumption of diet high in phytic acid with relatively high physical activity may be protective against the adverse effects of cadmium.     

Inhibition of neuronal nitric oxide synthase activity promotes migration of human‐induced pluripotent stem cell‐derived neural stem cells toward cancer cells

The breakthrough in derivation of human‐induced pluripotent stem cells (hiPSCs) provides an approach that may help overcome ethical and allergenic challenges posed in numerous medical applications involving human cells, including neural stem/progenitor cells (NSCs). Considering the great potential of NSCs in targeted cancer gene therapy, we investigated in this study the tumor tropism of hiPSC‐derived NSCs and attempted to enhance the tropism by manipulation of biological activities of proteins that are involved in regulating the migration of NSCs toward cancer cells. We first demonstrated that hiPSC‐NSCs displayed tropism for both glioblastoma cells and breast cancer cells in vitro and in vivo. We then compared gene expression profiles between migratory and non‐migratory hiPSC‐NSCs toward these cancer cells and observed that the gene encoding neuronal nitric oxide synthase (nNOS) was down‐regulated in migratory hiPSC‐NSCs. Using nNOS inhibitors and nNOS siRNAs, we demonstrated that this protein is a relevant regulator in controlling migration of hiPSC‐NSCs toward cancer cells, and that inhibition of its activity or down‐regulation of its expression can sensitize poorly migratory NSCs and be used to improve their tumor tropism. These findings suggest a novel application of nNOS inhibitors in neural stem cell‐mediated cancer therapy.     

Bivariate and Multivariate Analyses of the Influence of Blood Variables of Patients Submitted to Roux-en-Y Gastric Bypass on the Stability of Erythrocyte Membrane against the Chaotropic Action of Ethanol

The stability of the erythrocyte membrane, which is essential for the maintenance of cell functions, occurs in a critical region of fluidity, which depends largely on its composition and the composition and characteristics of the medium. As the composition of the erythrocyte membrane is influenced by several blood variables, the stability of the erythrocyte membrane must have relations with them. The present study aimed to evaluate, by bivariate and multivariate statistical analyses, the correlations and causal relationships between hematologic and biochemical variables and the stability of the erythrocyte membrane against the chaotropic action of ethanol. The validity of this type of analysis depends on the homogeneity of the population and on the variability of the studied parameters, conditions that can be filled by patients who undergo bariatric surgery by the technique of Roux-en-Y gastric bypass since they will suffer feeding restrictions that have great impact on their blood composition. Pathway analysis revealed that an increase in hemoglobin leads to decreased stability of the cell, probably through a process mediated by an increase in mean corpuscular volume. Furthermore, an increase in the mean corpuscular hemoglobin (MCH) leads to an increase in erythrocyte membrane stability, probably because higher values of MCH are associated with smaller quantities of red blood cells and a larger contact area between the cell membrane and ethanol present in the medium.     

Atypical Cohesin-Dockerin Complex Responsible for Cell Surface Attachment of Cellulosomal Components: BINDING FIDELITY,PROMISCUITY, AND STRUCTURAL BUTTRESSES*

The rumen bacterium Ruminococcus flavefaciens produces a highly organized multienzyme cellulosome complex that plays a key role in the degradation of plant cell wall polysaccharides, notably cellulose. The R. flavefaciens cellulosomal system is anchored to the bacterial cell wall through a relatively small ScaE scaffoldin subunit, which bears a single type IIIe cohesin responsible for the attachment of two major dockerin-containing scaffoldin proteins, ScaB and the cellulose-binding protein CttA. Although ScaB recruits the catalytic machinery onto the complex, CttA mediates attachment of the bacterial substrate via its two putative carbohydrate-binding modules. In an effort to understand the structural basis for assembly and cell surface attachment of the cellulosome in R. flavefaciens, we determined the crystal structure of the high affinity complex (K_d = 20.83 nm) between the cohesin module of ScaE (CohE) and its cognate X-dockerin (XDoc) modular dyad from CttA at 1.97-Å resolution. The structure reveals an atypical calcium-binding loop containing a 13-residue insert. The results further pinpoint two charged specificity-related residues on the surface of the cohesin module that are responsible for specific versus promiscuous cross-strain binding of the dockerin module. In addition, a combined functional role for the three enigmatic dockerin inserts was established whereby these extraneous segments serve as structural buttresses that reinforce the stalklike conformation of the X-module, thus segregating its tethered complement of cellulosomal components from the cell surface. The novel structure of the RfCohE-XDoc complex sheds light on divergent dockerin structure and function and provides insight into the specificity features of the type IIIe cohesin-dockerin interaction.     

Identification of Regions Important for Resistance and Signalling within the Antimicrobial Peptide Transporter BceAB of Bacillus subtilis

In the low-G+C-content Gram-positive bacteria, resistance to antimicrobial peptides is often mediated by so-called resistance modules. These consist of a two-component system and an ATP-binding cassette transporter and are characterized by an unusual mode of signal transduction where the transporter acts as a sensor of antimicrobial peptides, because the histidine kinase alone cannot detect the substrates directly. Thus, the transporters fulfill a dual function as sensors and detoxification systems to confer resistance, but the mechanistic details of these processes are unknown. The paradigm and best-understood example for this is the BceRS-BceAB module of Bacillus subtilis, which mediates resistance to bacitracin, mersacidin, and actagardine. Using a random mutagenesis approach, we here show that mutations that affect specific functions of the transporter BceAB are primarily found in the C-terminal region of the permease, BceB, particularly in the eighth transmembrane helix. Further, we show that while signaling and resistance are functionally interconnected, several mutations could be identified that strongly affected one activity of the transporter but had only minor effects on the other. Thus, a partial genetic separation of the two properties could be achieved by single amino acid replacements, providing first insights into the signaling mechanism of these unusual modules.