Large-scale antibody immune response mapping of splenic B cells and bone marrow plasma cells in a transgenic mouse model

Molecular characterization of antibody immunity and human antibody discovery is mainly carried out using peripheral memory B cells, and occasionally plasmablasts, that express B cell receptors (BCRs) on their cell surface. Despite the importance of plasma cells (PCs) as the dominant source of circulating antibodies in serum, PCs are rarely utilized because they do not express surface BCRs and cannot be analyzed using antigen-based fluorescence-activated cell sorting. Here, we studied the antibodies encoded by the entire mature B cell populations, including PCs, and compared the antibody repertoires of bone marrow and spleen compartments elicited by immunization in a human immunoglobulin transgenic mouse strain. To circumvent prior technical limitations for analysis of plasma cells, we applied single-cell antibody heavy and light chain gene capture from the entire mature B cell repertoires followed by yeast display functional analysis using a cytokine as a model immunogen. We performed affinity-based sorting of antibody yeast display libraries and large-scale next-generation sequencing analyses to follow antibody lineage performance, with experimental validation of 76 monoclonal antibodies against the cytokine antigen that identified three antibodies with exquisite double-digit picomolar binding affinity. We observed that spleen B cell populations generated higher affinity antibodies compared to bone marrow PCs and that antigen-specific splenic B cells had higher average levels of somatic hypermutation. A degree of clonal overlap was also observed between bone marrow and spleen antibody repertoires, indicating common origins of certain clones across lymphoid compartments. These data demonstrate a new capacity to functionally analyze antigen-specific B cell populations of different lymphoid organs, including PCs, for high-affinity antibody discovery and detailed fundamental studies of antibody immunity

Remobilization of leaf S compounds and senescence in response to restricted sulphate supply during the vegetative stage of oilseed rape are affected by mineral N availability

The impact of sulphur limitation on the remobilization of endogenous S compounds during the rosette stage of oilseed rape, and the interactions with N availability on these processes, were examined using a long-term 34SO42− labelling method combined with a study of leaf senescence progression (using SAG12/Cab as a molecular indicator) and gene expression of the transporters, BnSultr4;1 and BnSultr4;2, involved in vacuolar sulphate efflux. After 51 d on hydroponic culture at 0.3 mM 34SO42− (1 atom% excess), the labelling was stopped and plants were subject for 28 d to High S-High N (HS-HN, control), Low S-High N (LS-HN) or Low S-Low N (LS-LN) conditions. Compared with the control, LS-HN plants showed delayed leaf senescence and, whilst the shoot growth and the foliar soluble protein amounts were not affected, S, 34S, and SO42− amounts in the old leaves declined rapidly and were associated with the up-regulation of BnSultr4;1. In LS-LN plants, shoot growth was reduced, leaf senescence was accelerated, and the rapid S mobilization in old leaves was accompanied by decreased 34S and SO42−, higher protein mobilization, and up-regulation of BnSultr4;2, but without any change of expression of BnSultr4;1. The data suggest that to sustain the S demand for growth under S restriction (i) vacuolar SO42− is specifically remobilized in LS-HN conditions without any acceleration of leaf senescence, (ii) SO42− mobilization is related to an up-regulation of BnSultr4;1 and/or BnSultr4;2 expression, and (iii) the relationship between sulphate mobilization and up-regulation of expression of BnSultr4 genes is specifically dependent on the N availability

Combined use of modified atmosphere packaging and high pressure to extend the shelf-life of raw poultry sausage

International audienceThe contribution of modified atmosphere packaging (MAP) in extending the shelf-life of high-pressure treated raw poultry sausages was examined by considering microbial and oxidative stability (TBARs) aspects. Raw poultry sausages packaged under air or modified atmosphere (50% CO2–50% N2) were pressurized at 500 MPa during 5 min at a maximum temperature of 10.5 °C, subsequently allowed to refrigerated storage during 22 days. During storage, samples were tested at time intervals for headspace gas composition, pH, TBARs, Aerobic Mesophilic Counts (AMC) and Lactic Acid Bacteria (LAB) counts. The high pressure treatment could represent an efficient means of extending the microbiological shelf-life, insofar as it reduced and stabilized the AMC and LAB counts. However, the MAP did not further improve the microbial quality. But, still, by limiting lipid oxidation, it remains an essential technology for the control of the organoleptic quality, another important characteristic to consider in shelf-life determinations.Industrial relevanceRaw poultry meat and especially raw ground poultry meat, such as raw poultry sausages are highly perishable.Economic challenges and busier lifestyles have consumers seeking out products with longer and longer shelf-lives. Manufacturers have to respond to this demand by improving processes. The use of alternative preservation techniques such as high hydrostatic pressure represents a promising strategy to enhance the shelf-life of meat products and is preferred by consumers to addition of food additives. Modified atmosphere packaging is largely used to extend the shelf-life of processed meat products. Manufacturers may question the relevance of maintaining MAP while introducing a new step of high pressure treatment in their process. This study showed that MAP significantly reduced lipid oxidation. In that way, MAP remains necessary to maintain organoleptic quality of pressurized raw poultry sausages

EffectorK, a comprehensive resource to mine for Ralstonia, Xanthomonas,

International audiencePathogens deploy effector proteins that interact with host proteins to manipulate the host physiology to the pathogen's own benefit. However, effectors can also be recognized by host immune proteins, leading to the activation of defence responses. Effectors are thus essential components in determining the outcome of plant-pathogen interactions. Despite major efforts to decipher effector functions, our current knowledge on effector biology is scattered and often limited. In this study, we conducted two systematic large-scale yeast two-hybrid screenings to detect interactions betweenArabidopsis thalianaproteins and effectors from two vascular bacterial pathogens:Ralstonia pseudosolanacearumandXanthomonas campestris. We then constructed an interactomic network focused onArabidopsisand effector proteins from a wide variety of bacterial, oomycete, fungal, and invertebrate pathogens. This network contains our experimental data and protein-protein interactions from 2,035 peer-reviewed publications (48,200Arabidopsis-Arabidopsisand 1,300Arabidopsis-effector protein interactions). Our results show that effectors from different species interact with both common and specificArabidopsisinteractors, suggesting dual roles as modulators of generic and adaptive host processes. Network analyses revealed that effector interactors, particularly "effector hubs" and bacterial core effector interactors, occupy important positions for network organization, as shown by their larger number of protein interactions and centrality. These interactomic data were incorporated in EffectorK, a new graph-oriented knowledge database that allows users to navigate the network, search for homology, or find possible paths between host and/or effector proteins. EffectorK is available at and allows users to submit their own interactomic data