Molecular characterization of the Bacillus anthracis main S-layer component: evidence that it is the major cell-associated antigen

Bacillus anthracis, the aetiological agent of anthrax, is a Gram-positive spore-forming bacterium. The cell wall of vegetative cells of B. anthracis is surrounded by an S-layer. An array remained when sap, a gene described as encoding an S-layer component, was deleted. The remaining S-layer component, termed EA1, is chromosomally encoded. The gene encoding EA1 (eag) was obtained on two overlapping fragments in Escherichia coli and shown to be contiguous to the sap gene. The EA1 amino acid sequence, deduced from the eag nucleotide sequence, shows classical S-layer protein features (no cysteine, only 0.1% methionine, 10% lysine, and a weakly acidic pi). Similar to Sap and other Gram-positive surface proteins, EA1 has three 'S-layer-homology’motifs immediately downstream from a signal peptide. Single- and double-disrupted mutants were constructed. EA1 and Sap were co-localized at the cell surface of the wild-type bacilli. However, EA1 was more tightly bound than Sap to the bacteria. Electron microscopy studies and in vivo experiments with the constructed mutants showed that EA1 constitutes the main lattice of the B. anthracis S-layer, and is the major cell-associated antigen.     

Molecular characterization of the variable regions of a mouse polyreactive IgG2b antibody with rheumatoid factor activity

The complete nucleotide sequences of heavy and light chains of a mouse polyreactive IgG2b antibody were determined. This antibody, obtained after primary immunization of BALB/c mice with human lymphoblastoid cells, possess anti-HLA-DR and anti-rheumatoid factor activities and reacts with various self and nonself antigens. The VL and VH segments were found to belong to the VK8 and VH7183 families, respectively. The VH segment shared a high percentage of sequence similarity (95%) with previously described germline genes. The VK segment had 98.9% of sequence similarity with a consensus sequence VK8 of antibodies with anti-phosphorylcholine activity. Furthermore, the framework regions 2 and 3 of the VL segment were very similar to the framework regions 2 and 3 of other antibodies known to possess rheumatoid factor activity. We postulate that during immunization, the presence of HLA-DR antigens selects precursors having configurations similar to that of the germline, and induces some somatic mutations that do not significantly affect antibody polyreactivity.The nucleotide sequence data reported in this paper have been submitted to the EMBL nucleotide sequence database and have been assigned the accession numbers X53400 and X59816 for VH and VL respectively.     

Galactosyltransferase activities in subcellular fractions of two YC8 lymphoma cell variants: does a relationship exist between antigenic determinants and acceptor sites for galactose?

Experiments have been undertaken to correlate physiological changes, observed in two YC8 cells variants (P and L) and some of their immunological and enzymatic properties. These cell lines show different responses towards antilymphocyte and anti-Moloney sera. Subcellular fractionations have been made. The A fractions (d: 1.14/1.16) have the highest ouabain-inhibited Mg²⁺-stimulated (Na⁺-K⁺)-dependent ATPase and galactosyltransferase activities. Some properties of the latter enzyme have been studied: whereas optima pH and requirements for Mn²⁺ ions have been found to be the same for both cell line enzymes, on the contrary, different kinetic parameters have been shown with respect to sugar donor (UDP-galactose) on endogeneous or exogeneous (ovomucoid) acceptors. Apparent Km for UDP-galactose is 1.7 × 10⁻⁶ M (P-cells) and 3.3 × 10⁻⁶ M (L-cells), on endogeneous acceptors, and P-cell V max < L-cell V max; on ovomucoid it is 0.61 × 10⁻⁶ M, for both cell lines. These results suggest the presence on L-cells of more endogeneous acceptor sites, the higher affinity of P-cells for UDP-galactose being balanced by less endogeneous acceptor sites for galactose. When ovomucoid is added, galactose transfer on endogeneous acceptor sites of both cells is negligible. Apparent Km for ovomucoid is 8.6 × 10⁻⁵ M (P-cells) and 4.3 × 10⁻⁵ M (L-cells). These data support the above-mentioned hypothesis: L-cell enzymes would be more rapidly saturated than P-cell enzymes because of the higher number of endogeneous sites on L-cells.This supposed acquired character of L-cells as well as their immunological behaviour could explain the modified properties of L-cells as compared to P-cells.     

Us and them: From prejudice to racism. An original analysis of race and racism

This paper present the content of the Musée de l'Homme exhibition “Us and them: from prejudice to racism” and provides a detailed explanation of how this content has been presented to the public. In a second section of this paper we explain some of the analysis and concepts, from a biological anthropology perspective, that were the foundation of the exhibition and provides some information about the current situation in France.     

Influence of membrane-cortex linkers on the extrusion of membrane tubes

The cell membrane is an inhomogeneous system composed of phospholipids, sterols, carbohydrates, and proteins that can be directly attached to underlying cytoskeleton. The protein linkers between the membrane and the cytoskeleton are believed to have a profound effect on the mechanical properties of the cell membrane and its ability to reshape. Here, we investigate the role of membrane-cortex linkers on the extrusion of membrane tubes using computer simulations and experiments. In simulations, we find that the force for tube extrusion has a nonlinear dependence on the density of membrane-cortex attachments: at a range of low and intermediate linker densities, the force is not significantly influenced by the presence of the membrane-cortex attachments and resembles that of the bare membrane. For large concentrations of linkers, however, the force substantially increases compared with the bare membrane. In both cases, the linkers provided membrane tubes with increased stability against coalescence. We then pulled tubes from HEK cells using optical tweezers for varying expression levels of the membrane-cortex attachment protein Ezrin. In line with simulations, we observed that overexpression of Ezrin led to an increased extrusion force, while Ezrin depletion had a negligible effect on the force. Our results shed light on the importance of local protein rearrangements for membrane reshaping at nanoscopic scales.     

Impact of hyperthermic intraperitoneal chemotherapy on Hsp27 protein expression in serum of patients with peritoneal carcinomatosis

Despite the strong rationale for combining cytoreductive surgery (CRS) with hyperthermic intraperitoneal chemotherapy (HIPEC) in patients with peritoneal carcinomatosis, thermotolerance and chemoresistance might result from heat shock protein overexpression. The aim of the present study was thus to determine whether the heat shock protein 27 (Hsp27), a potential factor in resistance to treatment, could have a higher level in serum from patients under this combined therapy. Patients receiving CRS plus HIPEC for peritoneal carcinomatosis (group 1), patients with cancer or a history of cancer undergoing abdominal surgery (group 2), and patients without malignancies undergoing abdominal surgery (group 3) were included. Hsp27 serum levels were determined before and at different times following CRS and HIPEC using enzyme-linked immunosorbent assay. In group 1 (n = 25), the high Hsp27 levels, observed at the end of surgery compared with before (p < 0.0001), decreased during HIPEC, but remained significantly higher than before surgery (p < 0.0005). In groups 2 (n = 11) and 3 (n = 15), surgery did not significantly increase Hsp27 levels. A targeted molecular strategy, inhibiting Hsp27 expression in tumor tissue, could significantly reduce resistance to the combined CRS plus HIPEC treatment. This approach should be further assessed in a clinical phase I trial.     

Staphylococcus aureus FepA and FepB Proteins Drive Heme Iron Utilization in Escherichia coli

EfeUOB-like tripartite systems are widespread in bacteria and in many cases they are encoded by genes organized into iron-regulated operons. They consist of: EfeU, a protein similar to the yeast iron permease Ftrp1; EfeO, an extracytoplasmic protein of unknown function and EfeB, also an extracytoplasmic protein with heme peroxidase activity, belonging to the DyP family. Many bacterial EfeUOB systems have been implicated in iron uptake, but a prefential iron source remains undetermined. Nevertheless, in the case of Escherichia coli, the EfeUOB system has been shown to recognize heme and to allow extracytoplasmic heme iron extraction via a deferrochelation reaction. Given the high level of sequence conservations between EfeUOB orthologs, we hypothesized that heme might be the physiological iron substrate for the other orthologous systems. To test this hypothesis, we undertook characterization of the Staphylococcus aureus FepABC system. Results presented here indicate: i) that the S. aureus FepB protein binds both heme and PPIX with high affinity, like EfeB, the E. coli ortholog; ii) that it has low peroxidase activity, comparable to that of EfeB; iii) that both FepA and FepB drive heme iron utilization, and both are required for this activity and iv) that the E. coli FepA ortholog (EfeO) cannot replace FepA in FepB-driven iron release from heme indicating protein specificity in these activities. Our results show that the function in heme iron extraction is conserved in the two orthologous systems.