Compound A, a dissociated glucocorticoid receptor modulator, inhibits T-bet (Th1) and induces GATA-3 (Th2) activity in immune cells

Background

Compound A (CpdA) is a dissociating non-steroidal glucocorticoid receptor (GR) ligand which has anti-inflammatory properties exerted by down-modulating proinflammatory gene expression. By favouring GR monomer formation, CpdA does not enhance glucocorticoid (GC) response element-driven gene expression, resulting in a reduced side effect profile as compared to GCs. Considering the importance of Th1/Th2 balance in the final outcome of immune and inflammatory responses, we analyzed how selective GR modulation differentially regulates the activity of T-bet and GATA-3, master drivers of Th1 and Th2 differentiation, respectively.

Results

Using Western analysis and reporter gene assays, we show in murine T cells that, similar to GCs, CpdA inhibits T-bet activity via a transrepressive mechanism. Different from GCs, CpdA induces GATA-3 activity by p38 MAPK-induction of GATA-3 phosphorylation and nuclear translocation. CpdA effects are reversed by the GR antagonist RU38486, proving the involvement of GR in these actions. ELISA assays demonstrate that modulation of T-bet and GATA-3 impacts on cytokine production shown by a decrease in IFN-γ and an increase in IL-5 production, respectively.

Conclusions

Taken together, through their effect favoring Th2 over Th1 responses, particular dissociated GR ligands, for which CpdA represents a paradigm, hold potential for the application in Th1-mediated immune disorders.     

IL-17RA Signaling Reduces Inflammation and Mortality during Trypanosoma cruzi Infection by Recruiting Suppressive IL-10-Producing Neutrophils

Members of the IL-17 cytokine family play an important role in protection against pathogens through the induction of different effector mechanisms. We determined that IL-17A, IL-17E and IL-17F are produced during the acute phase of T. cruzi infection. Using IL-17RA knockout (KO) mice, we demonstrate that IL-17RA, the common receptor subunit for many IL-17 family members, is required for host resistance during T. cruzi infection. Furthermore, infected IL-17RA KO mice that lack of response to several IL-17 cytokines showed amplified inflammatory responses with exuberant IFN-γ and TNF production that promoted hepatic damage and mortality. Absence of IL-17RA during T. cruzi infection resulted in reduced CXCL1 and CXCL2 expression in spleen and liver and limited neutrophil recruitment. T. cruzi-stimulated neutrophils secreted IL-10 and showed an IL-10-dependent suppressive phenotype in vitro inhibiting T-cell proliferation and IFN-γ production. Specific depletion of Ly-6G+ neutrophils in vivo during T. cruzi infection raised parasitemia and serum IFN-γ concentration and resulted in increased liver pathology in WT mice and overwhelming wasting disease in IL-17RA KO mice. Adoptively transferred neutrophils were unable to migrate to tissues and to restore resistant phenotype in infected IL-17RA KO mice but migrated to spleen and liver of infected WT mice and downregulated IFN-γ production and increased survival in an IL-10 dependent manner. Our results underscore the role of IL-17RA in the modulation of IFN-γ-mediated inflammatory responses during infections and uncover a previously unrecognized regulatory mechanism that involves the IL-17RA-mediated recruitment of suppressive IL-10-producing neutrophils.     

GAC1, a gene encoding a putative GTPase-activating protein, regulates bikaverin biosynthesis in Fusarium verticillioides

Fusarium verticillioides (teleomorph Gibberella moniliformis) is an ascomycete known to produce a variety of secondary metabolites, including fumonisins, fusaric acid and bikaverin. These metabolites are synthesized when the fungus is under stress, notably nutrient limitations. To date we have limited understanding of the complex regulatory process associated with fungal secondary metabolism. In this study we generated a collection of F. verticillioides mutants by using REMI (restriction enzyme mediated integration) mutagenesis and in the process identified a strain, R647, that carries a mutation in a gene designated GAC1. Mutation in the GACI locus, which encodes a putative GTPase activating protein, resulted in the increased production of bikaverin, suggesting that GAC1 is negatively associated with bikaverin biosynthesis. Complementation of R647 with the wildtype GAC1 gene restored the bikaverin production level to that of the wild-type progenitor, demonstrating that gac1 mutation was directly responsible for the overproduction of bikaverin. We also demonstrated that AREA, encoding global nitrogen regulator, and PKS4, encoding polyketide synthase, are downstream genes that respectively are regulated positively and negatively by GAC1. Our results suggest that GAC1 plays an important role in signal transduction regulating bikaverin production in F. verticillioides.     

The functions of the "Greeting Ceremony" among male mantled howlers (Alouatta palliata) on Agaltepec Island, Mexico

Nonhuman primates use greeting behaviors as nonaggressive communicatory signals in multiple social contexts. Adult male mantled howlers (Alouatta palliata) perform a ritual greeting that has been associated with bond-strengthening functions. The aim of this study is to explore the greeting patterns of male howlers living on Agaltepec Island, Mexico. Specifically, we analyzed the relationships between greetings and several individual, relational, and contextual variables, such as the expression of affiliation and agonism, dominance rank, age, kinship relationships, spatial organization, activity patterns, and subgrouping patterns. Greetings were more frequent between males with closer dominance ranks. Among those dyads that greeted at least once, dominant males initiated greetings more frequently than less-dominant males. On the other hand, more greetings were observed when one of the participants had recently returned to a subgroup and during locomotion. On the basis of these results, we propose that on Agaltepec greetings are a conflict management mechanism used between males of similar ranks. The fission-fusion social system of this group of howlers allows males with conflicting interests to remain separated, and greetings may reduce tension during fusion events.     

Anaplasma marginale major surface protein 1a directs cell surface display of tick BM95 immunogenic peptides on Escherichia coli

The surface display of heterologous proteins on live Escherichia coli using anchoring motifs from outer membranes proteins has impacted on many areas of biochemistry, molecular biology and biotechnology. The Anaplasma marginale major surface protein 1a (MSP1a) contains N-terminal surface-exposed repeated peptides (28-289 amino acids) that are involved in pathogen interaction with host cell receptors and is surface-displayed when the recombinant protein is expressed in E. coli. Therefore, it was predicted that MSP1a would surface display on E. coli peptides inserted in the N-terminal repeats region of the protein. The Rhipicephalus (Boophilus) microplus BM86 and BM95 glycoproteins are homologous proteins that protect cattle against tick infestations. In this study, we demonstrated that a recombinant protein comprising tick BM95 immunogenic peptides fused to the A. marginale MSP1a N-terminal region is displayed on the E. coli surface and is recognized by anti-BM86 and anti-MSP1a antibodies. This system provides a novel approach to the surface display of heterologous antigenic proteins on live E. coli and suggests the possibility to use the recombinant bacteria for immunization studies against cattle tick infestations.     

NMR investigations of protein-carbohydrate interactions: insights into the topology of the bound conformation of a lactose isomer and beta-galactosyl xyloses to mistletoe lectin and galectin-1.

A hallmark of oligosaccharides is their often limited spatial flexibility, allowing them to access a distinct set of conformers in solution. Viewing each individual or even the complete ensemble of conformations as potential binding partner(s) for lectins in protein-carbohydrate interactions, it is pertinent to address the question on the characteristics of bound state conformation(s) in solution. Also, it is possible that entering the lectin's binding site distorts the low-energy topology of a glycosidic linkage. As a step to delineate the strategy of ligand selection for galactosides, a common physiological docking point, we have performed a NMR study on two non-homologous lectins showing identical monosaccharide specificity. Thus, the conformation of lactose analogues bound to bovine heart galectin-1 and to mistletoe lectin in solution has been determined by transferred nuclear Overhauser effect measurements. It is demonstrated that the lectins select the syn conformation of lactose and various structural analogues (Galbeta(1-->4)Xyl, Galbeta(1-->3)Xyl, Galbeta(1-->2)Xyl, and Galbeta(1-->3)Glc) from the ensemble of presented conformations. No evidence for conformational distortion was obtained. Docking of the analogues to the modeled binding sites furnishes explanations, in structural terms, for exclusive recognition of the syn conformer despite the non-homologous design of the binding sites.     

Nucleotide ester-forming alcoholytic activities of nucleotide pyrophosphatases: implications for practical biotransformation, enzyme mechanisms and biological function

Nucleotide pyrophosphatases (NPP) hydrolyze phosphoanhydride and phosphodiester derivatives of nucleoside 5′-monophosphates (NMP) yielding NMP as a product. In a water–alcohol mixture, the alcohol (R–OH) competes and substitutes for water as the splitting agent, so a mixture of NMP and NMP-O-alkyl ester (NMP-O-R) is formed. NPPs from snake venom, potato tuber and mammalian tissues have been studied in this regard. Snake and potato NPPs were considered as possible practical biocatalysts to synthesize NMP-O-Rs from various nucleotidic substrates and alcohols. Mammalian NPPs, mainly from human blood and rat liver, were studied considering the possibility that the alcoholytic reactions catalyzed by them could be biologically relevant. Valuable information on the active centers and catalytic mechanisms of NPPs was also obtained.     

A histochemical and immunohistochemical study of digestive enzymes and hormones during the larval development of the sea bream, Sparus aurata L.

Summary The distribution of different hydrolytic enzymes and the localization of the hormones which regulate glucose metabolism during development of the digestive tract of the sea bream, Sparus aurata L., were studied. The yolk sac contains trypsin, glucose-6-phosphatase, ATPases and acid and alkaline phosphatase activities. Positive insulin, glucagon and somatostatin cells were observed in the pancreas and in the lumen of the intestinal tract during endogenous feeding. From hatching until 3 days later, the digestive tract of sea bream larvae shows no enzymatic activities. During exogenous feeding, the activities of the phosphatases and trypsin generally increase, as do the amounts of the hydrolytic enzymes and trypsin, as well as the pancreatic and intestinal hormones. The enzymatic activities gradually decrease from the anterior part towards the posterior part of the digestive tract.     

Variation of bagasse crystallinity and cellulase activity during the fermentation of Cellulomonas bacteria

A characteristic behavior of the fermentation process was observed during the growth of Cellulomonas on sugarcane bagasse. At the early stage of the fermentation the crystallinity index of bagasse increased, suggesting that the major metabolized fraction corresponded to the hemicellulose during this stage. Some time later the crystallinity achieved a steady state and then deceased, which indicated that the most complex structure of bagasse was being attacked. The analysis of the cellulolytic activity of extracellular enzyme in the medium showed a sharp increase followed by an abrupt leveling off and decline in activity. These results along with the reduction of crystallinity index and bagasse utilization (70%) justify the assumption that the C₁ component was present in the cellulase complex synthesized by the bacteria.