Antibody specific for the peptide.major histocompatibility complex. Is it T cell receptor-like?

Antibodies recognizing peptide bound to a major histocompatibility complex (MHC) protein usually have a higher affinity for the composite peptide.MHC (pMHC) ligand than T cell receptors (TCR) with the same specificity. Because the solvent-accessible peptide area constitutes only a small portion of the contacting pMHC surface, we hypothesized that the contribution of the MHC moiety to the TCR-pMHC complex stability is limited, ensuring a small increment of the binding energy delivered by the peptide to be distinguishable by the TCR or the peptide-specific antibody. This suggests that the gain in affinity of the antibody-pMHC interaction can be achieved through an increase in the on-rate without a significant change in the off-rate of the interaction. To test the hypothesis, we have analyzed the binding of an ovalbumin peptide (pOV8) and its variants associated with soluble H-2Kb protein to the 25-D1.16 monoclonal antibody and compared it with the binding of the same pMHC complexes to the OT-1 TCR. This comparison revealed a substantially higher on-rate of the antibody-pMHC interaction compared with the TCR-pMHC interaction. In contrast, both the antibody and the TCR-pMHC complexes exhibited comparably fast off-rates. Sequencing of the 25-D1.16 VH and VL genes showed that they have very few somatic mutations and those occur mainly in framework regions. We propose that the above features constitute a signature of the recognition of MHC-bound peptide antigens by TCR and TCR-like antibodies, which could explain why the latter are rarely produced in vivo.     

Integrins Influence the Size and Dynamics of Signaling Microclusters in a Pyk2-dependent Manner

Integrin engagement on lymphocytes initiates “outside-in” signaling that is required for cytoskeleton remodeling and the formation of the synaptic interface. However, the mechanism by which the “outside-in” signal contributes to receptor-mediated intracellular signaling that regulates the kinetics of granule delivery and efficiency of cytolytic activity is not well understood. We have found that variations in ICAM-1 expression on tumor cells influence killing kinetics of these cells by CD16.NK-92 cytolytic effectors suggesting that changes in integrin ligation on the effector cells regulate the kinetics of cytolytic activity by the effector cells. To understand how variations of the integrin receptor ligation may alter cytolytic activity of CD16.NK-92 cells, we analyzed molecular events at the contact area of these cells exposed to planar lipid bilayers that display integrin ligands at different densities and activating CD16-specific antibodies. Changes in the extent of integrin ligation on CD16.NK-92 cells at the cell/bilayer interface revealed that the integrin signal influences the size and the dynamics of activating receptor microclusters in a Pyk2-dependent manner. Integrin-mediated changes of the intracellular signaling significantly affected the kinetics of degranulation of CD16.NK-92 cells providing evidence that integrins regulate the rate of target cell destruction in antibody-dependent cell cytotoxicity (ADCC).     

Production of soluble form of human TNF-alpha ligand-binding domain type 1 receptor by expression in Drosophila cells

5His-tagged human TNFalpha type I receptor (TNFR1) ligand-binding domain was produced in Drosophila cells under control of metallothionein Cu-inducible promoter and purified by Ni-NTA affinity chromatography to homogeneity. TNFR1 gene fragment was cloned by PCR from CD8+ in vitro cultured T-killer normal linage cDNA. In despite of three disulfide bonds, the recombinant protein was correctly folded which was conformed by TNFalpha ligand binding assay in ELISA variant.     

A new role for PGRP-S (Tag7) in immune defense: lymphocyte migration is induced by a chemoattractant complex of Tag7 with Mts1

PGRP-S (Tag7) is an innate immunity protein involved in the antimicrobial defense systems, both in insects and in mammals. We have previously shown that Tag7 specifically interacts with several proteins, including Hsp70 and the calcium binding protein S100A4 (Mts1), providing a number of novel cellular functions. Here we show that Tag7–Mts1 complex causes chemotactic migration of lymphocytes, with NK cells being a preferred target. Cells of either innate immunity (neutrophils and monocytes) or acquired immunity (CD4⁺ and CD8⁺ lymphocytes) can produce this complex, which confirms the close connection between components of the 2 branches of immune response.     

Effect of surface-active agents on peroxidase activity. IV. Effect of lipids and fatty acids from milk

The effect of surfactants, lipids and fatty acid salts isolated from cow milk on the activity of heme-containing horseradish peroxidase in solution was studied. As the surfactant concentration increases, the rate of the enzymic reaction successively decreases, increases, and again decreases, down to zero in the case of the fatty acid salts. The initial deceleration of the reaction rate results from the enzyme inhibition. The subsequent increase is caused by an improved accessibility for the substrate and the enhanced activity of the catalytic site of the enzyme due to its immobilization in the surfactant aggregates. A shielding of the protein by these aggregates can explain the secondary deceleration of the enzymic reaction rate. The general character of the dependence is similar and does not depend on the surfactant structure for a series of fatty acid salts and phospholipids; however, it is quite different in the case of cholesterol and sphingomyelin.     

Soft-walled monothalamids (Rhizaria: foraminifera) of the Crimean shelf (Black Sea): taxonomic composition and inter-regional patterns of species diversity and distribution

We present new data on monothalamous (single-chambered) foraminifera from the Black Sea Crimean shelf zone between Karkinitsky Gulf in the west to the area near Kerch in the east. Within this region we recognized a total of 40 morphospecies; 8 are assigned, in some cases tentatively, to known species and another 9 to known genera, again sometimes tentatively. Twelve species (all undescribed) are reported here for the first time. The most abundant species are Psammophaga sp. (sensu Gooday et al. 2011), Goodayia rostellata Sergeeva & Anikeeva 2008 and Vellaria pellucida Gooday & Fernando 1992, each of which is evenly distributed in all studied areas. The highest species richness of monothalamous foraminifera was observed in the Yalta region. Based on a multivariate analysis of monothalamid assemblage structure and diversity indices [Shannon (H’), Margalef (D’), eveness Pielou (J’), Simpson (1-λ’) and rarefaction ES(n)], we recognized three groups of stations, corresponding to the Western, Southern and Eastern coasts of the Crimean peninsula. The monothalamid assemblages found in each of these coastal regions exhibit different structural features and are distinguished by certain characteristic species.     

Efficient killing of tumor cells by CAR-T cells requires greater number of engaged CARs than TCRs

Although CAR-T cells are widely used to treat cancer, efficiency of CAR-T cell cytolytic responses has not been carefully examined. We engineered CAR specific for HMW-MAA (high-molecular-weight melanoma-associated antigen) and evaluated potency of CD8+ CAR-T cells to release cytolytic granules and to kill tissue-derived melanoma cells, which express different levels of HMW-MAA. CAR-T cells efficiently killed melanoma cells expressing high level of HMW-MAA, but not melanoma cells with lower levels of HMW-MAA. The same melanoma cells presenting significantly lower level of stimulatory peptide-MHC ligand were readily lysed by T cells transduced with genes encoding α,β-TCR specific for the peptide-MHC ligand. The data suggest that higher level of targeted molecules is required to engage a larger number of CARs than TCRs to induce efficient cytolytic granule release and destruction of melanoma cells. Understanding the difference in molecular mechanisms controlling activation thresholds of CAR- versus TCR-mediated responses will contribute to improving efficiency of CAR T cells required to eliminate solid tumors presenting low levels of targeted molecules.