The 3A6-TCR/superagonist/HLA-DR2a complex shows similar interface and reduced flexibility compared to the complex with self-peptide

T-cell receptor (TCR) recognition of the myelin basic protein (MBP) peptide presented by major histocompatibility complex (MHC) protein HLA-DR2a, one of the MHC class II alleles associated with multiple sclerosis, is highly variable. Interactions in the trimolecular complex between the TCR of the MBP83-99-specific T cell clone 3A6 with the MBP-peptide/HLA-DR2a (abbreviated TCR/pMHC) lead to substantially different proliferative responses when comparing the wild-type decapeptide MBP90-99 and a superagonist peptide, which differs mainly in the residues that point toward the TCR. Here, we investigate the influence of the peptide sequence on the interface and intrinsic plasticity of the TCR/pMHC trimolecular and pMHC bimolecular complexes by molecular dynamics simulations. The intermolecular contacts at the TCR/pMHC interface are similar for the complexes with the superagonist and the MBP self-peptide. The orientation angle between TCR and pMHC fluctuates less in the complex with the superagonist peptide. Thus, the higher structural stability of the TCR/pMHC tripartite complex with the superagonist peptide, rather than a major difference in binding mode with respect to the self-peptide, seems to be responsible for the stronger proliferative response.     

Intercellular compartmentation of basic carbon pathways in motor organs (pulvini) of leaves of Phaseolus coccineus L.

The activities of enzymes which catalyze one step in each of the five major carbon pathways in green plants were measured in secondary pulvini and other tissues of Phaseolus coccineus L. leaves. We were able to detect activities of fumarase (EC 4.2.1.2; tricarboxylic-acid pathway), NAD-glyceraldehyde-phosphate dehydrogenase (NAD-GAPDH, EC 1.2.1.12; glycolysis), 6-phosphogluconate dehydrogenase (6-PGDH, EC 1.1.1.44; oxidative pentose-phosphate pathway), ribulose-1, 5-bisphosphate carboxylase (Rubisco, EC 4.1.1.39; photosynthetic carbon-reduction pathway), and of hydroxypyruvate reductase (HP-R, EC 1.1.1.81; photosynthetic carbon-oxidation pathway). On a protein basis the activities of Rubisco and HP-R in pulvinar regions were very low (below 1 and 2 mol · (kg protein) ^–-1 · h^–-1, respectively), but the activities of fumarase and NAD-GAPDH were between 10- and 5-fold higher compared with the laminar tissue (up to 7 and 50 mol · (kg protein)^–-1 · h^–-1, respectively). Similarly, the protein specific activities of 6-PGDH were increased in the pulvinus (3–4 compared with approx. 1 mol · (kg protein)^–-1 · h^–-1 in the leaf blade). No differences in specific activities were detected between day and night positions of the leaves. By applying quantitative histochemical techniques we determined the longitudinal and transversal compartmentation of the activities of fumarase, NAD-GAPDH, and 6-PGDH in pulvinar tissues. Levels of activity of all three enzymes increased towards the middle part of the pulvinus. Here, expressed on a dry-weight (DW) basis, the analysis of cross sections showed highest activities in the outer parts of the extensor in the order given, approx. 0.6, 5, and 0.25 mol · (kg DW)^–-1 · h^–-1 for fumarase, NAD-GAPDH and 6-PGDH. When related to protein, levels of activity were comparably high within the inner parts of extensor and flexor, and partly also in the abaxial part of the bundle (fumarase, 6-PGDH). The tissue-specific compartmentation of the respective activities is discussed in relation to leaf movement and shows parallels with guard-cell function.Abbreviations Chl chlorophyll - DW dry weight - GAPDH glyceraldehyde-phosphate dehydrogenase - HP-R hydroxypyruvate reductase - Rubisco ribulose-1,5-bisphosphate carboxylase - 6-PGDH 6-phosphogluconate dehydrogenase This investigation was supported by a grant from the Deutsche Forschungsgemeinschaft.     

Transformation: how do nematode sperm become activated and crawl?

Nematode sperm undergo a drastic physiological change during spermiogenesis (sperm activation). Unlike mammalian flagellated sperm, nematode sperm are amoeboid cells and their motility is driven by the dynamics of a cytoskeleton composed of major sperm protein (MSP) rather than actin found in other crawling cells. This review focuses on sperm from Caenorhabditis elegans and Ascaris suum to address the roles of external and internal factors that trigger sperm activation and power sperm motility. Nematode sperm can be activated in vitro by several factors, including Pronase and ionophores, and in vivo through the TRY-5 and SPE-8 pathways. Moreover, protease and protease inhibitors are crucial regulators of sperm maturation. MSP-based sperm motility involves a coupled process of protrusion and retraction, both of which have been reconstituted in vitro. Sperm motility is mediated by phosphorylation signals, as illustrated by identification of several key components (MPOP, MFPs and MPAK) in Ascaris and the characterization of GSP-3/4 in C. elegans.     

Functional analysis of Leishmania major cyclophilin

A potent immunosuppressive drug cyclosporin A (CsA) is known to inhibit human cell infection by the pathogenic protozoan parasite Leishmania major both in vitro and in vivo. The proposed mechanism of action involves CsA binding to Leishmania major-expressed cyclophilin and subsequent down-regulation of signaling events necessary for establishing productive infection. Recently, we identified a ubiquitously expressed membrane protein, CD147, as a signaling receptor for extracellular cyclophilins in mammalian cells. Here we demonstrate that, while being enzymatically active, the Leishmania cyclophilin, unlike its human homologue, does not interact with CD147 on the cell surface of target cells. CD147 facilitates neither Leishmania binding nor infection. Primary structure and biochemical analyses revealed that the parasite's cyclophilin is defective in heparan binding, an event required for signaling interaction between CD147 and human cyclophilin. When the heparan-binding motif was reconstituted in Leishmania cyclophilin, it regained the CD147-dependent signaling activity. These results underscore a critical role of cyclophilin-heparan interactions in CD147-mediated signaling events and argue against the role of Leishmania cyclophilin in parasite binding to target cells.     

Expression and function of TETRAN, a new type of membrane transporter

In contrast to transport across basolateral membranes, the mechanism governing transport of organic anions across the luminal membranes of proximal tubules has remained unclear. We recently found Tetracycline transporter-like protein (TETRAN), a human ortholog of yeast Tpo1p that can transport anionic Non-steroidal anti-inflammatory drugs (NSAIDs). In this study, we examine the expression and function of TETRAN. TETRAN mRNA is expressed in various human tissues, including kidney. When overexpressed in cultured cells, TETRAN was predominantly localized on cytoplasmic membranes. Immunohistochemical analysis of human and mouse kidney tissue showed that TETRAN was expressed at the luminal membranes of proximal tubules. Overexpression of TETRAN in cultured cells facilitated the uptake of organic anions such as indomethacin (a NSAID) and fluorescein. The results suggest that TETRAN is a novel human organic anion transporter, and that it serves as a transporter for some NSAIDs and various other organic anions at the final excretion step.     

Leishmania infantum: provision of reducing equivalents to the mitochondrial tryparedoxin/tryparedoxin peroxidase system

Within the mitochondrion of Leishmania infantum, hydroperoxide metabolism relies on the activity of tryparedoxin-dependent peroxidases (TXNPxs). Tryparedoxins (TXNs) are thioredoxin-related oxidoreductases, which in vitro are reduced by the trypanothione reductase/trypanothione [TR/T(SH)₂] redox couple. Still, there is no evidence that this actually occurs in the mitochondrion. This communication addresses the question of how the mitochondrial TXN/TXNPx system is reduced. First, using a digitonin fractionation assay, we show that TR activity is absent from the L. infantum mitochondrion. The possibility that this organelle possesses alternative electron sources for TXN/TXNPx is then investigated. Biochemical assays performed with purified recombinant enzymes, revealed that TR and T(SH)₂ can be replaced, albeit less efficiently, by the dihydrolipoamide dehydrogenase/lipoamide redox system as TXN/TXNPx electron donor. This result challenges the classical view that T(SH)₂ is the only reductant for TXNs and add new prospects regarding the involvement of 2-oxo acid dehydrogenase complexes in L. infantum mitochondrial hydroperoxide metabolism.     

Leishmania chagasi: homogenous metacyclic promastigotes isolated by buoyant density are highly virulent in a mouse model

Homogenous metacyclic promastigotes of Leishmania chagasi were isolated by buoyant density from in vitro heterogeneous cultures and used for biochemical characterization of isoforms of the major surface protease (MSP). Compared to stationary phase promastigotes, metacyclic cells had three times more MSP, produced 3-fold higher parasite loads in a mouse model in vivo, and were more resistant to complement-mediated lysis in vitro. These metacyclic L. chagasi expressed both the virulence-associated 59-kDa, and the constitutively expressed 63-kDa, isoforms of MSP.     

Differential effects of a parasite on ornamental structures based on melanins and carotenoids

Models of sexual selection predict that ornamental colorationshould be affected by parasites in order to serve as honestsignals. Animals are commonly infested by a range of parasitespecies and often simultaneously display several ornaments.Thus the specific effect of a given parasite on ornaments isimportant for the understanding of the signal. Here we investigateexperimentally the effect of an ectoparasite on carotenoid-and melanin-based traits in breeding great tits Parus major.In the experiment, nests were either infested with hen fleas,Ceratophyllus gallinae, or kept free of parasites. The colorof the two traits and the size of the melanin-based breaststripe were assessed both in the year of experimental parasiteinfestation and during the following breeding season, afterthe annual molt. The size of the breast stripe of infested malesand females significantly decreased, but increased significantlyin uninfested males and females. The blackness of the breaststripe and the carotenoid-based plumage coloration was unaffected.Our experiment demonstrates that the size of the melanin-basedbreast stripe of adults depends on parasite infestation, suggestingthat the trait can serve as an honest signal of previous parasite exposure.     

Genome-wide analysis reveals increased levels of transcripts related with infectivity in peanut lectin non-agglutinated promastigotes of Leishmania infantum

Metacyclic promastigotes are transmitted during bloodmeals after development inside the gut of the sandfly vector. The isolation from axenic cultures of procyclic and metacyclic promastigotes by peanut lectin agglutination followed by differential centrifugation is controversial in Leishmania infantum. The purpose of this study has been to isolate both fractions simultaneously from the same population in stationary phase of axenic culture and compare their expression profiles by whole-genome shotgun DNA microarrays. The 317 genes found with meaningful values of stage-specific regulation demonstrate that negative selection of metacyclic promastigotes by PNA agglutination is feasible in L. infantum and both fractions can be isolated. This subpopulation up-regulates a cysteine peptidase A and several genes involved in lipophosphoglycan, proteophosphoglycan and glycoprotein biosynthesis, all related with infectivity. In fact, we have confirmed the increased infection rate of PNA⁻ promastigotes by U937 human cell line infection experiments. These data support that metacyclic promastigotes are related with infectivity and the lack of agglutination with PNA is a phenotypic marker for this subpopulation.     

MHC standing genetic variation and pathogen resistance in wild Atlantic salmon

Pathogens are increasingly emerging in human-altered environments as a serious threat to biodiversity. In this context of rapid environmental changes, improving our knowledge on the interaction between ecology and evolution is critical. The objective of this study was to evaluate the influence of an immunocompetence gene, the major histocompatibility complex (MHC) class IIβ, on the pathogen infection levels in wild Atlantic salmon populations, Salmo salar, and identify selective agents involved in contemporary coevolution. MHC variability and bacterial infection rate were determined throughout the summer in juvenile salmon from six rivers belonging to different genetic and ecological regions in Québec, Canada. A total of 13 different pathogens were identified in kidney by DNA sequence analysis, including a predominant myxozoa, most probably recently introduced in North America. Infection rates were the highest in southern rivers at the beginning of the summer (average 47.6±6.3% infected fish). One MHC allele conferred a 2.9 times greater chance of being resistant to myxozoa, while another allele increased susceptibility by 3.4 times. The decrease in frequency of the susceptibility allele but not other MHC or microsatellite alleles during summer was suggestive of a mortality event from myxozoa infection. These results supported the hypothesis of pathogen-driven selection in the wild by means of frequency-dependent selection or change in selection through time and space rather than heterozygous advantage, and underline the importance of MHC standing genetic variation for facing pathogens in a changing environment.