Probing the interaction between recombinant human myelin basic protein and caseins using surface plasmon resonance and diffusing wave spectroscopy

An intrinsically unstructured human myelin basic protein (hMBP) was expressed in the milk of transgenic cows (TGmilk) and found exclusively associated with the casein micellar phase. The interaction between the recombinant protein and milk caseins was investigated using surface plasmon resonance (SPR). An anti‐human myelin basic protein antibody was covalently immobilized to the surface of the sensor chip. Subsequently the interaction between the recombinant protein (captured by this antibody) and caseins was studied in comparison to that noted with its human counterpart. Results showed a calcium‐mediated interaction between the recombinant protein and caseins. The order of magnitude of this interaction was in agreement with the number of phosphorylated residues carried by each type of casein (α_s‐ > β‐ > κ‐casein). This selective interaction was not noted between the human protein and milk caseins indicating that the recombinant protein was phosphorylated to a higher extent than the human protein. The obtained results indicated that the co‐expression of the recombinant protein and caseins by the mammary gland along with the recombinant protein's ability to form calcium bridges played a key role in the association of the recombinant human myelin basic protein (rhMBP) with the casein micelles of milk. Despite this association between the recombinant protein and milk caseins, light scattering investigations using diffusing wave spectroscopy (DWS) showed no significant differences between the milks of the transgenic and the non‐transgenic control cows, with respect to both the average micelle size and surface charges. This was attributed to the low expression levels of the recombinant protein in milk. Copyright © 2009 John Wiley & Sons, Ltd.     

Interactions between Nosema microspores and a neonicotinoid weaken honeybees (Apis mellifera)

Global pollinators, like honeybees, are declining in abundance and diversity, which can adversely affect natural ecosystems and agriculture. Therefore, we tested the current hypotheses describing honeybee losses as a multifactorial syndrome, by investigating integrative effects of an infectious organism and an insecticide on honeybee health. We demonstrated that the interaction between the microsporidia Nosema and a neonicotinoid (imidacloprid) significantly weakened honeybees. In the short term, the combination of both agents caused the highest individual mortality rates and energetic stress. By quantifying the strength of immunity at both the individual and social levels, we showed that neither the haemocyte number nor the phenoloxidase activity of individuals was affected by the different treatments. However, the activity of glucose oxidase, enabling bees to sterilize colony and brood food, was significantly decreased only by the combination of both factors compared with control, Nosema or imidacloprid groups, suggesting a synergistic interaction and in the long term a higher susceptibility of the colony to pathogens. This provides the first evidences that interaction between an infectious organism and a chemical can also threaten pollinators, interactions that are widely used to eliminate insect pests in integrative pest management.     

Notch resolves mixed neural identities in the zebrafish epiphysis

Manipulation of Notch activity alters neuronal subtype identity in vertebrate neuronal lineages. Nonetheless, it remains controversial whether Notch activity diversifies cell fate by regulating the timing of neurogenesis or acts directly in neuronal subtype specification. Here, we address the role of Notch in the zebrafish epiphysis, a simple structure containing only two neural subtypes: projection neurons and photoreceptors. Reducing the activity of the Notch pathway results in an excess of projection neurons at the expense of photoreceptors, as well as an increase in cells retaining a mixed identity. However, although forced activation of the pathway inhibits the projection neuron fate, it does not promote photoreceptor identity. As birthdating experiments show that projection neurons and photoreceptors are born simultaneously, Notch acts directly during neuronal specification rather than by controlling the timing of neurogenesis. Finally, our data suggest that two distinct signals are required for photoreceptor fate specification: one for the induction of the photoreceptor fate and the other, involving Notch, for the inhibition of projection neuron traits. We propose a novel model in which Notch resolves mixed neural identities by repressing an undesired genetic program.     

Population genetics reveals origin and number of founders in a biological invasion

Propagule pressure is considered the main determinant of success of biological invasions: when a large number of individuals are introduced into an area, the species is more likely to establish and become invasive. Nevertheless, precise data on propagule pressure exist only for a small sample of invasive species, usually voluntarily introduced. We studied the invasion of the American bullfrog, Rana catesbeiana, into Europe, a species that is considered a major cause of decline for native amphibians. For this major invader with scarce historical data, we used population genetics data (a partial sequence of the mitochondrial cytochrome b gene) to infer the invasion history and to estimate the number of founders of non-native populations. Based on differences between populations, at least six independent introductions from the native range occurred in Europe, followed by secondary translocations. Genetic diversity was strongly reduced in non-native populations, indicating a very strong bottleneck during colonization. We used simulations to estimate the precise number of founders and found that most non-native populations derive from less than six females. This capability of invasion from a very small number of propagules challenges usual management strategies; species with such ability should be identified at an early stage of introduction.     

Exploring the pteridophyte flora of the Eastern Afromontane biodiversity hotspot

In this review, we explore our current understanding of the fern and lycophyte diversity occurring in the Eastern Afromontane Biodiversity Hotspot (EABH). The review explores the species diversity of this region in the context of the Afromadagascan pteridophyte diversity based on an exhaustive species list assembled in the synopsis of Afromadagascan pteridophytes published by Roux in 2009. The list was updated by incorporating recent progress in our understanding of the taxonomy and phylogeny of these plants. Evidence for a distinct pteridophyte flora occurring in the East African mountain region was discovered using ordination and clustering analyses. This EABH floras shares links to other Afromadagascan pteridophyte floras such as the one in the tropical lowland forests of central and western Africa. These floras share the dominance of species that preferably occur in humid climates whereas other African pteridophyte floras tend to contain a higher proportion of xeric adapted ferns. The phylogenetic composition of the EABH pteridophyte flora was assessed by comparing global versus local proportion of orders, families, and genera. This analysis revealed distinct patterns that are partly caused by the radiation of Blotiella and Triplophyllum besides selective colonization of species pre-adapted to Afromadagascan climates. In situ speciation in the East African tropical mountains may have contributed to the global diversity of widespread genera such as Asplenium and Pteris. In summary, this is the first comprehensive attempt to assess the pteridophyte diversity of the East African mountains providing the framework for future studies on their conservation, ecology, and evolution.     

Water-seeking behavior in insects harboring hairworms: should the host collaborate?

We explored the idea that hosts infected with manipulative parasitesmight mitigate the costs of infection by collaborating withthe parasite rather than resisting it. Nematomorphs are usuallyconsidered to be manipulative parasites of arthropods becausethey cause hosts to seek an aquatic environment, which is neededby the adult parasite. We placed infected cricket hosts in situationsof forced noncompliance and compared some fitness parameters(life expectancy, gonad development, and reproductive behaviors)in noncompliant hosts and hosts allowed to express parasite-inducedbehavior. Compared to uninfected controls, reduced survivalwas observed in both males and females from the two categoriesof infected hosts, collaborative or not. A substantial proportionof collaborative females produced eggs or had developed ovarieswhile such phenomena were never observed among noncollaborativeones. Collaborative females retained a nymphal phenotype, butadult males nevertheless courted and produced spermatophoresto such females. However, collaborative females had difficultiesmounting males, taking spermatophores and/or ovipositing. Incontrast to females, all males were entirely castrated by theparasite regardless of their behavior, collaborative or not.Thus, bringing the parasite into water does not effectivelymitigate the costs of infection for the host.     

PilT is required for PI(3,4,5)P3-mediated crosstalk between Neisseria gonorrhoeae and epithelial cells

The retractile type IV pilus participates in a number of fundamental bacterial processes, including motility, DNA transformation, fruiting body formation and attachment to host cells. Retraction of the N. gonorrhoeae type IV pilus requires a functional pilT. Retraction generates substantial force on its substrate (> 100 pN per retraction event), and it has been speculated that epithelial cells sense and respond to these forces during infection. We provide evidence that piliated, Opa non-expressing Neisseria gonorrhoeae activates the stress-responsive PI-3 kinase/Akt (PKB) pathway in human epithelial cells, and activation is enhanced by a functional pilT. PI-3 kinase inhibitors wortmannin and LY294002 reduce cell entry by 81% and 50%, respectively, illustrating the importance of this cascade in bacterial invasion. PI-3 kinase and its direct downstream effectors [PI(3,4,5)P3] and Akt are concentrated in the cell cortex beneath adherent bacteria, particularly at the periphery of the bacterial microcolonies. Furthermore, [PI(3,4,5)P3] is translocated to the outer leaflet of the plasma membrane. Finally, we show that [PI(3,4,5)P3] stimulates microcolony formation and upregulates pilT expression in vitro. We conclude that N. gonorrhoeae activation of PI-3 kinase triggers the host cell to produce a lipid second messenger that influences bacterial behaviour.     

Importin beta negatively regulates nuclear membrane fusion and nuclear pore complex assembly

Assembly of a eukaryotic nucleus involves three distinct events: membrane recruitment, fusion to form a double nuclear membrane, and nuclear pore complex (NPC) assembly. We report that importin beta negatively regulates two of these events, membrane fusion and NPC assembly. When excess importin beta is added to a full Xenopus nuclear reconstitution reaction, vesicles are recruited to chromatin but their fusion is blocked. The importin beta down-regulation of membrane fusion is Ran-GTP reversible. Indeed, excess RanGTP (RanQ69L) alone stimulates excessive membrane fusion, leading to intranuclear membrane tubules and cytoplasmic annulate lamellae-like structures. We propose that a precise balance of importin beta to Ran is required to create a correct double nuclear membrane and simultaneously to repress undesirable fusion events. Interestingly, truncated importin beta 45-462 allows membrane fusion but produces nuclei lacking any NPCs. This reveals distinct importin beta-regulation of NPC assembly. Excess full-length importin beta and beta 45-462 act similarly when added to prefused nuclear intermediates, i.e., both block NPC assembly. The importin beta NPC block, which maps downstream of GTPgammaS and BAPTA-sensitive steps in NPC assembly, is reversible by cytosol. Remarkably, it is not reversible by 25 microM RanGTP, a concentration that easily reverses fusion inhibition. This report, using a full reconstitution system and natural chromatin substrates, significantly expands the repertoire of importin beta. Its roles now encompass negative regulation of two of the major events of nuclear assembly: membrane fusion and NPC assembly.