EphB signaling directs peripheral nerve regeneration through Sox2-dependent Schwann cell sorting.

The peripheral nervous system has astonishing regenerative capabilities in that cut nerves are able to reconnect and re-establish their function. Schwann cells are important players in this process, during which they dedifferentiate to a progenitor/stem cell and promote axonal regrowth. Here, we report that fibroblasts also play a key role. Upon nerve cut, ephrin-B/EphB2 signaling between fibroblasts and Schwann cells results in cell sorting, followed by directional collective cell migration of Schwann cells out of the nerve stumps to guide regrowing axons across the wound. Mechanistically, we find that cell-sorting downstream of EphB2 is mediated by the stemness factor Sox2 through N-cadherin relocalization to Schwann cell-cell contacts. In vivo, loss of EphB2 signaling impaired organized migration of Schwann cells, resulting in misdirected axonal regrowth. Our results identify a link between Ephs and Sox proteins, providing a mechanism by which progenitor cells can translate environmental cues to orchestrate the formation of new tissue.

Nanobody inhibitors of Plexin-B1 identify allostery in plexin-semaphorin interactions and signaling.

Plexin-B1 is a receptor for the cell surface semaphorin, Sema4D. This signaling system has been implicated in a variety of human diseases, including cancer, multiple sclerosis and osteoporosis. While inhibitors of the Plexin-B1:Sema4D interaction have been previously reported, understanding their mechanism has been hindered by an incomplete structural view of Plexin-B1. In this study, we have raised and characterized a pair of nanobodies that are specific for mouse Plexin-B1 and which inhibit the binding of Sema4D to mouse Plexin-B1 and its biological activity. Structural studies of these nanobodies reveal that they inhibit the binding of Sema4D in an allosteric manner, binding to epitopes not previously reported. In addition, we report the first unbound structure of human Plexin-B1, which reveals that Plexin-B1 undergoes a conformational change on Sema4D binding. These changes mirror those seen upon binding of allosteric peptide modulators, which suggests a new model for understanding Plexin-B1 signaling and provides a potential innovative route for therapeutic modulation of Plexin-B1.

An antagonistic monoclonal anti-Plexin-B1 antibody exerts therapeutic effects in mouse models of postmenopausal osteoporosis and multiple sclerosis.

Osteoporosis and multiple sclerosis are highly prevalent diseases with limited treatment options. In light of these unmet medical needs, novel therapeutic approaches are urgently sought. Previously, the activation of the transmembrane receptor Plexin-B1 by its ligand semaphorin 4D (Sema4D) has been shown to suppress bone formation and promote neuroinflammation in mice. However, it is unclear whether inhibition of this receptor-ligand interaction by an anti-Plexin-B1 antibody could represent a viable strategy against diseases related to these processes. Here, we raised and systematically characterized a monoclonal antibody directed against the extracellular domain of human Plexin-B1, which specifically blocks the binding of Sema4D to Plexin-B1. In vitro, we show that this antibody inhibits the suppressive effects of Sema4D on human osteoblast differentiation and mineralization. To test the therapeutic potential of the antibody in vivo, we generated a humanized mouse line, which expresses transgenic human Plexin-B1 instead of endogenous murine Plexin-B1. Employing these mice, we demonstrate that the anti-Plexin-B1 antibody exhibits beneficial effects in mouse models of postmenopausal osteoporosis and multiple sclerosis in vivo. In summary, our data identify an anti-Plexin-B1 antibody as a potential therapeutic agent for the treatment of osteoporosis and multiple sclerosis.

Bombali Virus in Mops condylurus Bats, Guinea.

In 2018, a previously unknown Ebola virus, Bombali virus, was discovered in Sierra Leone. We describe detection of Bombali virus in Guinea. We found viral RNA in internal organs of 3 Angolan free-tailed bats (Mops condylurus) trapped in the city of N'Zerekore and in a nearby village.

Genetic Structure of Aedes (Stegomyia) albopictus Populations in Russia.

Background: Aedes (Stegomyia) albopictus was found for the first time in 2011 on the Black Sea coast in Russia, and during 2011-2019, the species expanded over two climate zones Cfa and Csa. Methods: Here, we studied the sequence diversity of the mitochondrial cytochrome c oxidase I (COI) gene, 1317-1433bp in length. In total, 131 specimens of Ae. albopictus sampled from 21 locations in Russia and Abkhazia were examined. Results: Two of the six identified mitochondrial haplotypes were detected for the first time. Four COI haplotypes were shared by at least two studied local populations. The most prevalent H1 and H2 haplotypes dominated in all the sampled localities in the Cfa zone. The H3 haplotype was prevalent in the Csa zone. Other haplotypes were rare. Phylogenetic analyses, spatial isolation and limited gene flow revealed that the samples from the Csa zone differed significantly from those from the Cfa zone. Conclusion: Two spatially isolated genetic lineages exist in Ae. albopictus population in southern region of Russia. One lineage obtained on the seacoast and inland (in valleys of the Caucasus Mountains and steppe zone) is widely distributed worldwide including Mediterranean populations. This confirms the hypothesis that the emergence of Ae. albopictus population in southern region of Russia may be associated with the terrestrial spread of mosquitoes from the well-established European population due to human activity. The other lineage, discovered in Novorossiysk, a maritime port, is similar to Ae. albopictus from the USA and Japan, suggesting the independent introduction of these mosquitoes.

Detection of Filoviruses in Bats in Vietnam.

A new filovirus named Menglà virus was found in bats in southern China in 2015. This species has been assigned to the new genus Dianlovirus and has only been detected in China. In this article, we report the detection of filoviruses in bats captured in Vietnam. We studied 248 bats of 15 species caught in the provinces of Lai Chau and Son La in northern Vietnam and in the province of Dong Thap in the southern part of the country. Filovirus RNA was found in four Rousettus leschenaultii and one Rousettus amplexicaudatus from Lai Chau Province. Phylogenetic analysis of the polymerase gene fragment showed that three positive samples belong to Dianlovirus, and two samples form a separate clade closer to Orthomarburgvirus. An enzyme-linked immunosorbent assay showed that 9% of Rousettus, 13% of Eonycteris, and 10% of Cynopterus bats had antibodies to the glycoprotein of marburgviruses.

The division protein FtsZ interacts with the small heat shock protein IbpA in Acholeplasma laidlawii.

Small heat shock proteins (sHSPs) control the proteins stability in the cell preventing their irreversible denaturation. While many mycoplasmas possess the sHSP gene in the genome, Acholeplasma laidlawii is the only mycoplasma capable of surviving in the environment. Here we report that the sHSP IbpA directly interacts with the key division protein FtsZ in A. laidlawii, representing the first example of such interaction in prokaryotes. FtsZ co-immunoprecipitates with IbpA from A. laidlawii crude extract and in vitro binds IbpA with KD ~ 1 µM. Proteins co-localize in the soluble fraction of the cell at 30-37 °C and in the non-soluble fraction after 1 h exposition to cold stress (4 °C). Under heat shock conditions (42 °C) the amount of FtsZ decreases and the protein remains in both soluble and non-soluble fractions. Furthermore, in vitro, FtsZ co-elutes with IbpAHis6 from A. laidlawii crude extract at any temperatures from 4 to 42 °C, with highest yield at 42 °C. Moreover, in vitro FtsZ retains its GTPase activity in presence of IbpA, and the filaments and bundles formation seems to be even improved by sHSP at 30-37 °C. At extreme temperatures, either 4 or 42 °C, IbpA facilitates FtsZ polymerization, although filaments under 4 °C appears shorter and with lower density, while at 42 °C IbpA sticks around the bundles, preventing their destruction by heat. Taken together, these data suggest that sHSP IbpA in A. laidlawii contributes to the FtsZ stability control and may be assisting appropriate cell division under unfavorable conditions.

A semaphorin-plexin-Rasal1 signaling pathway inhibits gastrin expression and protects against peptic ulcers.

Peptic ulcer disease is a frequent clinical problem with potentially serious complications such as bleeding or perforation. A decisive factor in the pathogenesis of peptic ulcers is gastric acid, the secretion of which is controlled by the hormone gastrin released from gastric G cells. However, the molecular mechanisms regulating gastrin plasma concentrations are poorly understood. Here, we identified a semaphorin-plexin signaling pathway that operates in gastric G cells to inhibit gastrin expression on a transcriptional level, thereby limiting food-stimulated gastrin release and gastric acid secretion. Using a systematic siRNA screening approach combined with biochemical, cell biology, and in vivo mouse experiments, we found that the RasGAP protein Rasal1 is a central mediator of plexin signal transduction, which suppresses gastrin expression through inactivation of the small GTPase R-Ras. Moreover, we show that Rasal1 is pathophysiologically relevant for the pathogenesis of peptic ulcers induced by nonsteroidal anti-inflammatory drugs (NSAIDs), a main risk factor of peptic ulcers in humans. Last, we show that application of recombinant semaphorin 4D alleviates peptic ulcer disease in mice in vivo, demonstrating that this signaling pathway can be harnessed pharmacologically. This study unravels a mode of G cell regulation that is functionally important in gastric homeostasis and disease.

A tri-functional vanadium(iv) complex to detect cysteine oxidation.

The development of effective molecular probes to detect and image the levels of oxidative stress in cells remains a challenge. Herein we report the design, synthesis and preliminary biological evaluation of a novel optical probe to monitor oxidation of thiol groups in cysteine-based phosphatases (CBPs). Following orthogonal protecting approaches we synthesised a new vanadyl complex designed to bind to CBPs. This complex is functionalised with a well-known dimedone derivative (to covalently trap sulfenic acids, SOHs) and a coumarin-based fluorophore for optical visualization. We show that this new probe efficiently binds to a range of phosphatases in vitro with nanomolar affinity. Moreover, preliminary flow cytometry and microscopy studies in live HCT116 cells show that this probe can successfully image cellular levels of sulfenic acids - one of the species resulting from protein oxidative damage.

Vanadyl complexes with dansyl-labelled di-picolinic acid ligands: synthesis, phosphatase inhibition activity and cellular uptake studies.

Vanadium complexes have been previously utilised as potent inhibitors of cysteine based phosphatases (CBPs). Herein, we present the synthesis and characterisation of two new fluorescently labelled vanadyl complexes (14 and 15) with bridged di-picolinic acid ligands. These compounds differ significantly from previous vanadyl complexes with phosphatase inhibition properties in that the metal-chelating part is a single tetradentate unit, which should afford greater stability and scope for synthetic elaboration than the earlier complexes. These new complexes inhibit a selection of cysteine based phosphatases (CBPs) in the nM range with some selectivity. Fluorescence spectroscopic studies (including fluorescence anisotropy) were carried out to demonstrate that the complexes are not simply acting as vanadyl delivery vehicles but they interact with the proteins. Finally, we present preliminary fluorescence microscopy studies to demonstrate that the complexes are cell permeable and localise throughout the cytoplasm of NIH3T3 cells.

Purification, quantification, and functional analysis of Complement Factor H.

Complement Factor H (FH) is an abundant, non-enzymic plasma/serum glycoprotein, which has a major role in regulating activation of the complement system. It can be purified from human plasma/serum by affinity chromatography, using a monoclonal anti-FH antibody as ligand. Other affinity chromatography ligands, including cardiolipin and trinitrophenyl-bovine serum albumin (TNP-BSA), can be used to purify human FH and also FH from a wide range of vertebrates, including mammals, birds, bony fish. Human FH protein concentration can be quantified by sandwich ELISA. The activity of FH is generally measured by assays which detect the cleavage, by complement factor I, of the complement protein C3b to form iC3b. Cleavage occurs only in the presence of a cofactor, and FH is one of a small number of cofactors for this reaction.

Comparative RNAi screening identifies a conserved core metazoan actinome by phenotype.

Although a large number of actin-binding proteins and their regulators have been identified through classical approaches, gaps in our knowledge remain. Here, we used genome-wide RNA interference as a systematic method to define metazoan actin regulators based on visual phenotype. Using comparative screens in cultured Drosophila and human cells, we generated phenotypic profiles for annotated actin regulators together with proteins bearing predicted actin-binding domains. These phenotypic clusters for the known metazoan "actinome" were used to identify putative new core actin regulators, together with a number of genes with conserved but poorly studied roles in the regulation of the actin cytoskeleton, several of which we studied in detail. This work suggests that although our search for new components of the core actin machinery is nearing saturation, regulation at the level of nuclear actin export, RNA splicing, ubiquitination, and other upstream processes remains an important but unexplored frontier of actin biology.

Epidemiology of West Nile infection in Volgograd, Russia, in relation to climate change and mosquito (Diptera: Culicidae) bionomics.

In 1999, there was the large outbreak of West Nile fever (WNF) in Southern Russia (>500 cases in the Volgograd Province). In 2000-2004, the WNF incidence rate decreased steadily to zero, but a new outbreak occurred in 2007 (64 cases). The analysis of historical climate data for Volgograd from 1900 to present showed that the years 1999 and 2007 were the hottest ones due to a very mild "winter" (Dec.-Mar.) and a hot "summer" (June-Sep.). There are up to 15 potential WNF vectors in Volgograd, but only Culex pipiens and Culex modestus are abundant in late summer, both in urban and rural settings. Only these species are naturally attracted to and feed on both humans and birds. The RNA of pathogenic WN virus genovariant was found by reverse transcriptase polymerase chain reaction only in Culex mosquitoes at the infection rate of about 0.04%. So these species may be considered as potential WNF "bridge vectors" between birds and humans as well as main vectors in sylvatic avain cycle. Their abundance in an epidemic season was higher in the years with a mild winter and a hot summer, so this phenomenon may serve as a connecting link between a climate and WNF epidemiology. These findings give some hints on the predisposing factors for WNF epidemic as well as the possibility to predict WNF outbreaks in the temperate climate zones.