Suppression of TNF-α and IL-1 signaling identifies a mechanism of homeostatic regulation of macrophages by IL-27

IL-27 is a pleiotropic cytokine with both activating and inhibitory functions on innate and acquired immunity. IL-27 is expressed at sites of inflammation in cytokine-driven autoimmune/inflammatory diseases, such as rheumatoid arthritis, psoriasis, inflammatory bowel disease, and sarcoidosis. However, its role in modulating disease pathogenesis is still unknown. In this study, we found that IL-27 production is induced by TNF-α in human macrophages (MΦ) and investigated the effects of IL-27 on the responses of primary human MΦ to the endogenous inflammatory cytokines TNF-α and IL-1. In striking contrast to IL-27-mediated augmentation of TLR-induced cytokine production, we found that IL-27 suppressed MΦ responses to TNF-α and IL-1β, thus identifying an anti-inflammatory function of IL-27. IL-27 blocked the proximal steps of TNF-α signaling by downregulating cell-surface expression of the signaling receptors p55 and p75. The mechanism of inhibition of IL-1 signaling was downregulation of the ligand-binding IL-1RI concomitant with increased expression of the receptor antagonist IL-1Ra and the decoy receptor IL-1RII. These findings provide a mechanism for suppressive effects of IL-27 on innate immune cells and suggest that IL-27 regulates inflammation by limiting activation of MΦ by inflammatory cytokines while preserving initial steps in host defense by augmenting responses to microbial products.     

Why do zoos attract crows? A comparative study from Europe and Asia

Crows have successfully colonized many cities, and urban zoos have been important in this process. To evaluate why zoos attract crows, we quantified crow numbers and behavior in three zoos in Europe (Debrecen, Edinburgh, Vienna) and one in Asia (Sapporo). Data were collected in 445 surveys over 297 days in summer 2014 and winter 2014–2015. We found that crow numbers were highest in Vienna, intermediate in Debrecen and Edinburgh and lowest in Sapporo, increased significantly from summer to winter (Debrecen, Edinburgh, Vienna), and from mornings to afternoons (Debrecen, Sapporo, Vienna), and were higher in sunny weather than in cloudy weather with precipitation and when visitor numbers were low (Debrecen, Vienna). The crows' use of natural food was highest in Vienna, intermediate in Edinburgh and Sapporo, and low in Debrecen. The use of anthropogenic food was high in Debrecen and Sapporo, where the availability of open grassy areas typically used by crows for natural foraging was low. In Sapporo, food availability was more limited than in other zoos, resulting in strong territoriality and few crows in summer, which decreased further in winter. Our study indicates that crows are primarily attracted to zoos by food availability and secondarily by breeding opportunities and that the relative importance of natural versus anthropogenic food sources may vary with zoo habitat structure. Our study draws attention to a previously overlooked role of zoos in urban biodiversity conservation. It may also provide useful information for the management of crow populations, if necessary, and for the planning of urban areas.     

An evaluation of electrophoresis as a taxonomic method using comparative data from the macropodidae (Marsupialia)

Comparative electrophoretic data collected for seven proteins from thirty-four species of marsupial have been examined to define the advantages and shortcomings of electrophoresis as a taxonomic method. The problems of relative rates of change, convergence and parallelism are examined and the range of taxonomic ranks which can usefully be studied with this technique is determined. The relative merits of this type of data, when combined with various methods of taxonomic analysis, are discussed.     

Break in the heat capacity change at 303 K for complex binding of netropsin to AATT containing hairpin DNA constructs

Studies performed in our laboratory demonstrated the formation of two thermodynamically distinct complexes on binding of netropsin to a number of hairpin-forming DNA sequences containing AATT-binding regions. These two complexes were proposed to differ only by a bridging water molecule between the drug and the DNA in the lower affinity complex. A temperature-dependent isothermal titration calorimetry (ITC)-binding study was performed using one of these constructs (a 20-mer hairpin of sequence 5'-CGAATTCGTCTCCGAATTCG) and netropsin. This study demonstrated a break in the heat capacity change for the formation of the complex containing the bridging water molecule at approximately 303 K. In the plot of the binding enthalpy change versus temperature, the slope (DeltaCp) was -0.67 kcal mol-1 K-1 steeper after the break at 303 K. Because of the relatively low melting temperature of the 20-mer hairpin (341 K (68 degrees C)), the enthalpy change for complex formation might have included some energy of refolding of the partially denatured hairpin, giving the suggestion of a larger DeltaCp. Studies done on the binding of netropsin to similar constructs, a 24-mer and a 28-mer, with added GC basepairs in the hairpin stem to increase thermal stability, exhibit the same nonlinearity in DeltaCp over the temperature range of from 275 to 333 K. The slopes (DeltaCp) were -0.69 and -0.64 kcal mol-1 K-1 steeper after 303 K for the 24-mer and 28-mer, respectively. This observation strengthens the argument regarding the presence of a bridging water molecule in the lower affinity netropsin/DNA complex. The DeltaCp data seem to infer that because the break in the heat capacity change function for the lower affinity binding occurs at the isoequilibrium temperature for water, water may be included or trapped in the complex. The fact that this break does not occur in the heat capacity change function for formation of the higher affinity complex can similarly be taken as evidence that water is not included in the higher affinity complex.     

Novel haem co-ordination variants of flavocytochrome P450BM3

Bacillus megaterium flavocytochrome P450 BM3 is a catalytically self-sufficient fatty acid hydroxylase formed by fusion of soluble NADPH-cytochrome P450 reductase and P450 domains. Selected mutations at residue 264 in the haem (P450) domain of the enzyme lead to novel amino acid sixth (distal) co-ordination ligands to the haem iron. The catalytic, spectroscopic and thermodynamic properties of the A264M, A264Q and A264C variants were determined in both the intact flavocytochromes and haem domains of P450 BM3. Crystal structures of the mutant haem domains demonstrate axial ligation of P450 haem iron by methionine and glutamine ligands trans to the cysteine thiolate, creating novel haem iron ligand sets in the A264M/Q variants. In contrast, the crystal structure of the A264C variant reveals no direct interaction between the introduced cysteine side chain and the haem, although EPR data indicate Cys(264) interactions with haem iron in solution. The A264M haem potential is elevated by comparison with wild-type haem domain, and substrate binding to the A264Q haem domain results in a approximately 360 mV increase in potential. All mutant haem domains occupy the conformation adopted by the substrate-bound form of wild-type BM3, despite the absence of added substrate. The A264M mutant (which has higher dodecanoate affinity than wild-type BM3) co-purifies with a structurally resolved lipid. These data demonstrate that a single mutation at Ala(264) is enough to perturb the conformational equilibrium between substrate-free and substrate-bound P450 BM3, and provide firm structural and spectroscopic data for novel haem iron ligand sets unprecedented in nature.     

Termite mounds contain soil-derived methanotroph communities kinetically adapted to elevated methane concentrations

Termite mounds have recently been confirmed to mitigate approximately half of termite methane (CH₄) emissions, but the aerobic CH₄ oxidising bacteria (methanotrophs) responsible for this consumption have not been resolved. Here, we describe the abundance, composition and CH₄ oxidation kinetics of the methanotroph communities in the mounds of three distinct termite species sampled from Northern Australia. Results from three independent methods employed show that methanotrophs are rare members of microbial communities in termite mounds, with a comparable abundance but distinct composition to those of adjoining soil samples. Across all mounds, the most abundant and prevalent methane monooxygenase sequences were affiliated with upland soil cluster α (USCα), with sequences homologous to Methylocystis and tropical upland soil cluster (TUSC) also detected. The reconstruction of a metagenome-assembled genome of a mound USCα representative highlighted the metabolic capabilities of this group of methanotrophs. The apparent Michaelis–Menten kinetics of CH₄ oxidation in mounds were estimated from in situ reaction rates. Methane affinities of the communities were in the low micromolar range, which is one to two orders of magnitude higher than those of upland soils, but significantly lower than those measured in soils with a large CH₄ source such as landfill cover soils. The rate constant of CH₄ oxidation, as well as the porosity of the mound material, were significantly positively correlated with the abundance of methanotroph communities of termite mounds. We conclude that termite-derived CH₄ emissions have selected for distinct methanotroph communities that are kinetically adapted to elevated CH₄ concentrations. However, factors other than substrate concentration appear to limit methanotroph abundance and hence these bacteria only partially mitigate termite-derived CH₄ emissions. Our results also highlight the predominant role of USCα in an environment with elevated CH₄ concentrations and suggest a higher functional diversity within this group than previously recognised.Subject terms: Soil microbiology, Biogeochemistry