Rapid cloning of high-affinity human monoclonal antibodies against influenza virus

Pre-existing neutralizing antibody provides the first line of defence against pathogens in general. For influenza virus, annual vaccinations are given to maintain protective levels of antibody against the currently circulating strains. Here we report that after booster vaccination there was a rapid and robust influenza-specific IgG+ antibody-secreting plasma cell (ASC) response that peaked at approximately day 7 and accounted for up to 6% of peripheral blood B cells. These ASCs could be distinguished from influenza-specific IgG+ memory B cells that peaked 14-21 days after vaccination and averaged 1% of all B cells. Importantly, as much as 80% of ASCs purified at the peak of the response were influenza specific. This ASC response was characterized by a highly restricted B-cell receptor (BCR) repertoire that in some donors was dominated by only a few B-cell clones. This pauci-clonal response, however, showed extensive intraclonal diversification from accumulated somatic mutations. We used the immunoglobulin variable regions isolated from sorted single ASCs to produce over 50 human monoclonal antibodies (mAbs) that bound to the three influenza vaccine strains with high affinity. This strategy demonstrates that we can generate multiple high-affinity mAbs from humans within a month after vaccination. The panel of influenza-virus-specific human mAbs allowed us to address the issue of original antigenic sin (OAS): the phenomenon where the induced antibody shows higher affinity to a previously encountered influenza virus strain compared with the virus strain present in the vaccine. However, we found that most of the influenza-virus-specific mAbs showed the highest affinity for the current vaccine strain. Thus, OAS does not seem to be a common occurrence in normal, healthy adults receiving influenza vaccination.     

T cells become licensed in the lung to enter the central nervous system

The blood–brain barrier (BBB) and the environment of the central nervous system (CNS) guard the nervous tissue from peripheral immune cells. In the autoimmune disease multiple sclerosis, myelin-reactive T-cell blasts are thought to transgress the BBB and create a pro-inflammatory environment in the CNS, thereby making possible a second autoimmune attack that starts from the leptomeningeal vessels and progresses into the parenchyma. Using a Lewis rat model of experimental autoimmune encephalomyelitis, we show here that contrary to the expectations of this concept, T-cell blasts do not efficiently enter the CNS and are not required to prepare the BBB for immune-cell recruitment. Instead, intravenously transferred T-cell blasts gain the capacity to enter the CNS after residing transiently within the lung tissues. Inside the lung tissues, they move along and within the airways to bronchus-associated lymphoid tissues and lung-draining mediastinal lymph nodes before they enter the blood circulation from where they reach the CNS. Effector T cells transferred directly into the airways showed a similar migratory pattern and retained their full pathogenicity. On their way the T cells fundamentally reprogrammed their gene-expression profile, characterized by downregulation of their activation program and upregulation of cellular locomotion molecules together with chemokine and adhesion receptors. The adhesion receptors include ninjurin 1, which participates in T-cell intravascular crawling on cerebral blood vessels. We detected that the lung constitutes a niche not only for activated T cells but also for resting myelin-reactive memory T cells. After local stimulation in the lung, these cells strongly proliferate and, after assuming migratory properties, enter the CNS and induce paralytic disease. The lung could therefore contribute to the activation of potentially autoaggressive T cells and their transition to a migratory mode as a prerequisite to entering their target tissues and inducing autoimmune disease.     

Major Transglycosylation Productsof β-Glucosidase and Their Induction Effects on Cellulase Biosynthesis

以纤维二糖特异诱导培养里氏木霉和黑曲霉菌丝,制备含有细胞外、细胞内和细胞质膜结合态β葡萄糖苷酶的无细胞抽提物,采用色谱持谱联用分析,比较研究他们对纤维二糖的转糖基作用,结果表明两菌株中都只有细胞质膜结合态β葡萄糖苷酶表现出显著的转糖基活性,可检测到的含有β糖苷键葡二糖类的主要转糖基产物是β龙胆二糖而不是槐糖．当在里氏木霉和黑曲霉菌丝的洗脱置换培养体系中加入β龙胆二糖时,可以诱导纤维素酶合成,但它比槐糖（迄今最有潜力的诱导物）的诱导效率低．这些结果支持假说认为是质膜结合态β葡萄糖苷酶从纤维素寡聚物形成转糖基作用产物,作为纤维素酶合成的诱导物     

Does Rft1 flip an N-glycan lipid precursor?

Protein N-glycosylation requires flipping of the glycolipid Man(5)GlcNAc(2)-diphosphate dolichol (Man(5)GlcNAc(2)-PP-Dol) across the endoplasmic reticulum (ER). Helenius et al. report genetic evidence suggesting that Rft1, an essential ER membrane protein in yeast, is required directly to translocate Man(5)GlcNAc(2)-PP-Dol. We now show that a specific ER protein(s), but not Rft1, is required to flip Man(5)GlcNAc(2)-PP-Dol in reconstituted vesicles. Rft1 may have a critical accessory role in translocating Man(5)GlcNAc(2)-PP-Dol in vivo, but the Man(5)GlcNAc(2)-PP-Dol flippase itself remains to be identified.     

Human gut microbiome viewed across age and geography

Gut microbial communities represent one source of human genetic and metabolic diversity. To examine how gut microbiomes differ among human populations, here we characterize bacterial species in fecal samples from 531 individuals, plus the gene content of 110 of them. The cohort encompassed healthy children and adults from the Amazonas of Venezuela, rural Malawi and US metropolitan areas and included mono- and dizygotic twins. Shared features of the functional maturation of the gut microbiome were identified during the first three years of life in all three populations, including age-associated changes in the genes involved in vitamin biosynthesis and metabolism. Pronounced differences in bacterial assemblages and functional gene repertoires were noted between US residents and those in the other two countries. These distinctive features are evident in early infancy as well as adulthood. Our findings underscore the need to consider the microbiome when evaluating human development, nutritional needs, physiological variations and the impact of westernization.     

Intrinsic motions along an enzymatic reaction trajectory

The mechanisms by which enzymes achieve extraordinary rate acceleration and specificity have long been of key interest in biochemistry. It is generally recognized that substrate binding coupled to conformational changes of the substrate-enzyme complex aligns the reactive groups in an optimal environment for efficient chemistry. Although chemical mechanisms have been elucidated for many enzymes, the question of how enzymes achieve the catalytically competent state has only recently become approachable by experiment and computation. Here we show crystallographic evidence for conformational substates along the trajectory towards the catalytically competent 'closed' state in the ligand-free form of the enzyme adenylate kinase. Molecular dynamics simulations indicate that these partially closed conformations are sampled in nanoseconds, whereas nuclear magnetic resonance and single-molecule fluorescence resonance energy transfer reveal rare sampling of a fully closed conformation occurring on the microsecond-to-millisecond timescale. Thus, the larger-scale motions in substrate-free adenylate kinase are not random, but preferentially follow the pathways that create the configuration capable of proficient chemistry. Such preferred directionality, encoded in the fold, may contribute to catalysis in many enzymes.     

Cochlear Implant

1 Introduction1 1.1 About Hearing Mechanisms The hearing function in human beings is something very specific and difficult to understand because it uses the brain highest functions. Basically, we can say (figure 1) that several stages are involved [1]: It is not easy to give a unique interpretation to each one of these stages, as most of the involved processes overlap at all levels. Nevertheless, as a brief summary, let us assume that [2]: l The ear transmits the air vibrations and transfor…     

Additive threats from pathogens, climate and land-use change for global amphibian diversity

Amphibian population declines far exceed those of other vertebrate groups, with 30% of all species listed as threatened by the International Union for Conservation of Nature. The causes of these declines are a matter of continued research, but probably include climate change, land-use change and spread of the pathogenic fungal disease chytridiomycosis. Here we assess the spatial distribution and interactions of these primary threats in relation to the global distribution of amphibian species. We show that the greatest proportions of species negatively affected by climate change are projected to be found in Africa, parts of northern South America and the Andes. Regions with the highest projected impact of land-use and climate change coincide, but there is little spatial overlap with regions highly threatened by the fungal disease. Overall, the areas harbouring the richest amphibian faunas are disproportionately more affected by one or multiple threat factors than areas with low richness. Amphibian declines are likely to accelerate in the twenty-first century, because multiple drivers of extinction could jeopardize their populations more than previous, mono-causal, assessments have suggested.