Affinity measured by microcluster

Like T cell activation, B cell activation is driven by aggregation of B cell receptors (BCRs) into microclusters. New work suggests that the early dynamics of BCR mobility and microcluster formation “translate” BCR affinity for antigen into B cell responsiveness

Organic matter composition and heterotrophic bacterial activity at declining summer sea ice in the central Arctic Ocean

The Arctic Ocean is highly susceptible to climate change as evidenced by rapid warming and the drastic loss of sea ice during summer. The consequences of these environmental changes for the microbial cycling of organic matter are largely unexplored. Here, we investigated the distribution and composition of dissolved organic matter (DOM) along with heterotrophic bacterial activity in seawater and sea ice of the Eurasian Basin at the time of the record ice minimum in 2012. Bacteria in seawater were highly responsive to fresh organic matter and remineralized on average 55% of primary production in the upper mixed layer. Correlation analysis showed that the accumulation of dissolved combined carbohydrates (DCCHO) and dissolved amino acids (DAA), two major components of fresh organic matter, was related to the drawdown of nitrate. Nitrate‐depleted surface waters at stations adjacent to the Laptev Sea showed about 25% higher concentrations of DAA than stations adjacent to the Barents Sea and in the central Arctic basin. Carbohydrate concentration was the best predictor of heterotrophic bacterial activity in sea ice. In contrast, variability in sea‐ice bacterial biomass was largely driven by differences in ice thickness. This decoupling of bacterial biomass and activity may mitigate the negative effects of biomass loss due to ice melting on heterotrophic bacterial functions. Overall, our results reveal that changes in DOM production and inventories induced by sea‐ice loss have a high potential to enhance the bacterial remineralization of organic matter in seawater and sea ice of the Arctic Ocean

Direct Visualization of Peptide/MHC Complexes at the Surface and in the Intracellular Compartments of Cells Infected In Vivo by Leishmania major

Protozoa and bacteria infect various types of phagocytic cells including macrophages, monocytes, dendritic cells and eosinophils. However, it is not clear which of these cells process and present microbial antigens in vivo and in which cellular compartments parasite peptides are loaded onto Major Histocompatibility Complex molecules. To address these issues, we have infected susceptible BALB/c (H-2d) mice with a recombinant Leishmania major parasite expressing a fluorescent tracer. To directly visualize the antigen presenting cells that present parasite-derived peptides to CD4+ T cells, we have generated a monoclonal antibody that reacts to an antigenic peptide derived from the parasite LACK antigen bound to I-Ad Major Histocompatibility Complex class II molecule. Immunogold electron microscopic analysis of in vivo infected cells showed that intracellular I-Ad/LACK complexes were present in the membrane of amastigote-containing phagosomes in dendritic cells, eosinophils and macrophages/monocytes. In both dendritic cells and macrophages, these complexes were also present in smaller vesicles that did not contain amastigote. The presence of I-Ad/LACK complexes at the surface of dendritic cells, but neither on the plasma membrane of macrophages nor eosinophils was independently confirmed by flow cytometry and by incubating sorted phagocytes with highly sensitive LACK-specific hybridomas. Altogether, our results suggest that peptides derived from Leishmania proteins are loaded onto Major Histocompatibility Complex class II molecules in the phagosomes of infected phagocytes. Although these complexes are transported to the cell surface in dendritic cells, therefore allowing the stimulation of parasite-specific CD4+ T cells, this does not occur in other phagocytic cells. To our knowledge, this is the first study in which Major Histocompatibility Complex class II molecules bound to peptides derived from a parasite protein have been visualized within and at the surface of cells that were infected in vivo

Human Neonatal Dendritic Cells Are Competent in MHC Class I Antigen Processing and Presentation

Neonates are clearly more susceptible to severe disease following infection with a variety of pathogens than are adults. However, the causes for this are unclear and are often attributed to immunological immaturity. While several aspects of immunity differ between adults and neonates, the capacity of dendritic cells in neonates to process and present antigen to CD8+ T cells remains to be addressed. We used human CD8+ T cell clones to compare the ability of neonatal and adult monocyte-derived dendritic cells to present or process and present antigen using the MHC class I pathway. Specifically, we assessed the ability of dendritic cells to present antigenic peptide, present an HLA-E–restricted antigen, process and present an MHC class I-restricted antigen through the classical MHC class I pathway, and cross present cell-associated antigen via MHC class I. We found no defect in neonatal dendritic cells to perform any of these processing and presentation functions and conclude that the MHC class I antigen processing and presentation pathway is functional in neonatal dendritic cells and hence may not account for the diminished control of pathogens

Calcium Influx Rescues Adenylate Cyclase-Hemolysin from Rapid Cell Membrane Removal and Enables Phagocyte Permeabilization by Toxin Pores

Bordetella adenylate cyclase toxin-hemolysin (CyaA) penetrates the cytoplasmic membrane of phagocytes and employs two distinct conformers to exert its multiple activities. One conformer forms cation-selective pores that permeabilize phagocyte membrane for efflux of cytosolic potassium. The other conformer conducts extracellular calcium ions across cytoplasmic membrane of cells, relocates into lipid rafts, translocates the adenylate cyclase enzyme (AC) domain into cells and converts cytosolic ATP to cAMP. We show that the calcium-conducting activity of CyaA controls the path and kinetics of endocytic removal of toxin pores from phagocyte membrane. The enzymatically inactive but calcium-conducting CyaA-AC− toxoid was endocytosed via a clathrin-dependent pathway. In contrast, a doubly mutated (E570K+E581P) toxoid, unable to conduct Ca2+ into cells, was rapidly internalized by membrane macropinocytosis, unless rescued by Ca2+ influx promoted in trans by ionomycin or intact toxoid. Moreover, a fully pore-forming CyaA-ΔAC hemolysin failed to permeabilize phagocytes, unless endocytic removal of its pores from cell membrane was decelerated through Ca2+ influx promoted by molecules locked in a Ca2+-conducting conformation by the 3D1 antibody. Inhibition of endocytosis also enabled the native B. pertussis-produced CyaA to induce lysis of J774A.1 macrophages at concentrations starting from 100 ng/ml. Hence, by mediating calcium influx into cells, the translocating conformer of CyaA controls the removal of bystander toxin pores from phagocyte membrane. This triggers a positive feedback loop of exacerbated cell permeabilization, where the efflux of cellular potassium yields further decreased toxin pore removal from cell membrane and this further enhances cell permeabilization and potassium efflux

A Liposome-Based Mycobacterial Vaccine Induces Potent Adult and Neonatal Multifunctional T Cells through the Exquisite Targeting of Dendritic Cells

BACKGROUND: In the search for more potent and safer tuberculosis vaccines, CAF01 was identified as a remarkable formulation. Based on cationic liposomes and including a synthetic mycobacterial glycolipid as TLR-independent immunomodulator, it induces strong and protective T helper-1 and T helper-17 adult murine responses to Ag85B-ESAT-6, a major mycobacterial fusion protein. Here, we assessed whether these properties extend to early life and how CAF01 mediates its adjuvant properties in vivo. METHODS/FINDINGS: Following adult or neonatal murine immunization, Ag85B-ESAT-6/CAF01 similarly reduced the post-challenge bacterial growth of M. bovis BCG, whereas no protection was observed using Alum as control. This protection was mediated by the induction of similarly strong Th1 and Th17 responses in both age groups. Multifunctional Th1 cells were already elicited after a single vaccine dose and persisted at high levels for at least 6 months even after neonatal priming. Unexpectedly, this potent adjuvanticity was not mediated by a massive targeting/activation of dendritic cells: in contrast, very few DCs in the draining lymph nodes were bearing the labeled antigen/adjuvant. The increased expression of the CD40 and CD86 activation markers was restricted to the minute portion of adjuvant-bearing DCs. However, vaccine-associated activated DCs were recovered several days after immunization. CONCLUSION: The potent adult and neonatal adjuvanticity of CAF01 is associated in vivo with an exquisite but prolonged DC uptake and activation, fulfilling the preclinical requirements for novel tuberculosis vaccines to be used in early life

Differential responses of bacteria to diatom-derived dissolved organic matter in the Arctic Ocean

International audienc

Heteroclitic proliferative responses and changes in cytokine profile induced by altered peptides: Implications for autoimmunity

Productive engagement of T cell receptors (TCRs) by cognate ligand (major histocompatibility complex plus peptide) leads to proliferation, differentiation, and the elaboration of effector functions. Altered peptides generated by single amino acid substitutions in the antigenic peptide have diverse effects on the outcome of the T cell response. We have generated an altered peptide (Q144) from an autoantigenic peptide of myelin proteolipid protein 139–151 by a single amino acid substitution (from tryptophan to glutamine) in the primary TCR contact at position 144 that is capable of inducing CD4(+) T cell responses in H-2(s) mice. By using a Q144-specific T cell clone (Q1.1B6), we see a hierarchy in T cell proliferation and cytokine production with various position 144 substituted peptides and have identified a peptide (L144) that hyperstimulates this T cell clone. In contrast to Q144, L144 induces maximal proliferation at 7 logs lower antigen concentration, induces greater cell death at higher antigen dose, and induces the secretion of cytokines not detected following stimulation with the cognate ligand. This heteroclitic T cell response associated with changes in cytokine profile was observed with several other T cell clones of different specificities. The L144 peptide also induces costimulation independent proliferation and cytokine production from the Q1.1B6 T cell clone. We describe this as a superagonist response. Such responses may have a role in the initiation of autoimmunity by promoting a proinflammatory environment following ligation of a cross-reactive TCR on autoreactive T cells