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.     

Hormonal regulation of a chicken oviduct messenger ribonucleic acid that shares a common domain with gizzard myosin light chain kinase

Changes in myosin light chain kinase (MLCK) and calmodulin (CaM) mRNAs have been evaluated during estrogen-mediated differentiation of the chicken oviduct. Also examined were acute changes that occur in oviduct RNA from animals stimulated with estrogen, withdrawn from hormone and then injected for 1, 2, and 4 days with synthetic estrogen [diethylstilbestrol (DES)], progesterone (P), or testosterone (T). Small changes were noted in both CaM and MLCK RNAs during primary stimulation when oviduct cells are actively dividing. On the other hand no significant changes were observed during secondary stimulation regardless of the steroid hormone injected. These data support the contention that CaM and MLCK are constitutively expressed but vary as a function of cell cycle. The MLCK mRNA is 5.5 kilobases (kb) but the MLCK cDNA also hybridizes to an oviduct RNA 2.7 kb long. This RNA species is acutely regulated by estrogen, P, and T but in a manner different from that of ovalbumin mRNA. The magnitude of stimulation of the 2.7 kb mRNA by diethylstilbestrol and T is greater than that of ovalbumin whereas changes in response to P are similar. The 12- to 16-fold increase of the 2.7 kb mRNA in response to T is the largest effect reported for this hormone acting on oviduct. The 2.7 kb mRNA encodes an unknown protein yet contains a 520 nucleotide segment that is highly homologous with the COOH-terminal coding portion of the MLCK mRNA. Since this homology does not include either catalytic or CaM-binding domains of MLCK, it is unlikely that the 2.7 kb mRNA encodes a CaM-dependent protein kinase.(ABSTRACT TRUNCATED AT 250 WORDS)     

Functional domains of chicken gizzard myosin light chain kinase

The proteolytic susceptibility of chicken gizzard myosin light chain kinase, a calmodulin-dependent enzyme, has been utilized to define the relative location of the catalytic and regulatory domains of the enzyme. Myosin light chain kinase isolated from this source exhibits a Mr of 130,000 and is extremely sensitive to trypsin at 24 degrees C; however, the molecule is divided into susceptible and resistant domains such that proteolysis proceeds rapidly and at multiple sites in the sensitive regions even at 4 degrees C while the rest of the molecule remains relatively resistant to digestion. One of these sensitive areas is the calmodulin-binding domain. On the other hand, Staphylococcus aureus V8 protease digestion generates a calmodulin-binding fragment (Mr = 70,000) that retains Ca2+/calmodulin-dependent enzymatic activity and both of the phosphorylation sites recognized by cAMP-dependent protein kinase. In contrast, treatment with chymotrypsin produces a 95,000 Mr calmodulin-binding fragment that contains only the calmodulin-modulated phosphorylation site. Sequential proteolytic digestion studies demonstrated that the chymotryptic cleavage site responsible for the generation of this 95,000 Mr peptide is within 3,000 Mr of the V8 protease site which produces the 70,000 Mr fragment. Moreover, the non-calmodulin-modulated phosphorylation site must exist in this 3,000 Mr region. A calmodulin-Sepharose affinity adsorption protocol was developed for the digestion and used to isolate both the 70,000 and 95,000 Mr fragments for further study. Taken together, our results are compatible with a model for chicken gizzard myosin light chain kinase in which there is no overlap between the active site, the calmodulin-binding region, and the two sites phosphorylated by cAMP-dependent protein kinase with regard to their relative position in the primary sequence of the molecule.     

Identification in turkey gizzard of an acidic protein related to the C-terminal portion of smooth muscle myosin light chain kinase

The isolation of an acidic protein, pI 4.5, that is abundant in turkey gizzard is described. Its apparent molecular weight measured by electrophoretic procedures is 24,000. This protein is phosphorylated by the catalytic subunit of the cAMP-dependent protein kinase and one phosphorylation site is indicated. From sequence determinations of tryptic peptides it is concluded that this protein is closely related to the C-terminal part of smooth muscle myosin light chain kinase. The initiation site for the protein is to the C-terminal side of the calmodulin-binding site. From the sequence data an estimated molecular weight is 18,000. This protein is expressed independently, as indicated by a blocked N terminus, and is probably the translation product of the 2.7-kilobase RNA detected previously in chicken gizzard (Guerriero, V., Jr., Russo, M. A., Olson, N. J., Putkey, J. A., and Means, A. R. (1986) Biochemistry 25, 8372-8381). Because of its putative origin as the C-terminal end of smooth muscle myosin light chain kinase, it is termed "telokin" (from a combination of kinase and the Greek telos, "end").     

Monitoring tropical forests under a functional perspective with satellite‐based vegetation optical depth

Monitoring ecosystem functions in forests is a priority in a climate change scenario, as climate‐induced events may initially alter the functions more than slow‐changing attributes, such as biomass. The ecosystem functional properties (EFPs) are quantities that characterize key ecosystem processes. They can be derived by point observations of gas and energy exchanges between the ecosystems and the atmosphere that are collected globally at FLUXNET flux tower sites and upscaled at ecosystem level. The properties here considered describe the ability of ecosystems to optimize the use of resources for carbon uptake. They represent functional forest information, are dependent on environmental drivers, linked to leaf traits and forest structure, and influenced by climate change effects. The ability of vegetation optical depth (VOD) to provide forest functional information is investigated using 2011–2014 satellite data collected by the Soil Moisture and Ocean Salinity mission and using the EFPs as reference dataset. Tropical forests in Africa and South America were analyzed, also according to ecological homogeneous units. VOD jointly with water deficit information explained 93% and 87% of the yearly variability in both flux upscaled maximum gross primary productivity and light use efficiency functional properties, in Africa and South America forests respectively. Maps of the retrieved properties evidenced changes in forest functional responses linked to anomalous climate‐induced events during the study period. The findings indicate that VOD can support the flux upscaling process in the tropical range, affected by high uncertainty, and the detection of forest anomalous functional responses. Preliminary temporal analysis of VOD and EFP signals showed fine‐grained variability in periodicity, in signal dephasing, and in the strength of the relationships. In selected drier forest types, these satellite data could also support the monitoring of functional dynamics.     

Application of an optimized marker amplification protocol in immunohistochemistry — a valuable tool for visualizing antigenic sites of low signal strength or sparse distribution

Amplification of immunohistochemical markers received considerable attention during the 1980s and 1990s. The amplification approach was largely abandoned following the development of antigen retrieval and reporter amplification techniques, because the latter were incorporated more easily into high throughput automated procedures in industrial and diagnostic laboratories. There remain, however, a number of instances where marker amplification still has much to offer. Consequently, we examined experimentally the utility of an optimized marker amplification technique in diagnostically relevant tissue where either the original signal strength was low or positive sites were visible, but sparsely distributed. Marker amplification in the former case not only improved the visibility of existing positive sites, but also revealed additional sites that previously were undetectable. In the latter case, positive sites were rendered more intense and therefore more easily seen during low magnification examination of large areas of tissue.     

Sensitivity of Aspergillus nidulans to the Cellulose Synthase Inhibitor Dichlobenil: Insights from Wall-Related Genes’ Expression and Ultrastructural Hyphal Morphologies

The fungal cell wall constitutes an important target for the development of antifungal drugs, because of its central role in morphogenesis, development and determination of fungal-specific molecular features. Fungal walls are characterized by a network of interconnected glycoproteins and polysaccharides, namely α-, β-glucans and chitin. Cell walls promptly and dynamically respond to environmental stimuli by a signaling mechanism, which triggers, among other responses, modulations in wall biosynthetic genes’ expression. Despite the absence of cellulose in the wall of the model filamentous fungus Aspergillus nidulans, we found in this study that fungal growth, spore germination and morphology are affected by the addition of the cellulose synthase inhibitor dichlobenil. Expression analysis of selected genes putatively involved in cell wall biosynthesis, carried out at different time points of drug exposure (i.e. 0, 1, 3, 6 and 24 h), revealed increased expression for the putative mixed linkage β-1,3;1,4 glucan synthase celA together with the β-1,3-glucan synthase fksA and the Rho-related GTPase rhoA. We also compared these data with the response to Congo Red, a known plant/fungal drug affecting both chitin and cellulose biosynthesis. The two drugs exerted different effects at the cell wall level, as shown by gene expression analysis and the ultrastructural features observed through atomic force microscopy and scanning electron microscopy. Although the concentration of dichlobenil required to affect growth of A. nidulans is approximately 10-fold higher than that required to inhibit plant cellulose biosynthesis, our work for the first time demonstrates that a cellulose biosynthesis inhibitor affects fungal growth, changes fungal morphology and expression of genes connected to fungal cell wall biosynthesis.     

EFFECT OF VITAMIN E SUPPLEMENTATION AND PARTIAL SUBSTITUTION OF POLY- WITH MONO-UNSATURATED FATTY ACIDS IN PIG DIETS ON MUSCLE,AND MICROSOME EXTRACT a-TOCOPHEROL CONCENTRATION AND LIPID OXIDATION

The experiment was organized in a 3×2 factorial arrangement with three dietary fat blends and a basal (20 mg kg^?1 diet) or supplemented (220 mg kg^?1) level of α-tocopheryl acetate. Dietary vitamin E and monounsaturated to polyunsaturated fatty acid ratio (dietary MUFA/PUFA) affected muscle α-tocopherol concentration (α-tocopherol [log μg g^?1]=0.18 (±0.105)+0.0034 (±0.0003)·dietary α-tocopherol [mg kg^?1 diet] (P<0.0001)+0.39 (±0.122)·dietary MUFA/PUFA (P<0.0036)). An interaction between dietary α-tocopherol and dietary MUFA/PUFA exists for microsome α-tocopherol concentration (α-tocopherol [log μg g^?1]=1.14 (±0.169) (P<0.0001)+0.0056 (±0.00099)·dietary α-tocopherol [mg kg^?1 diet] (P<0.0001)+0.54 (±0.206)·dietary MUFA/PUFA (P<0.0131)?0.0033 (±0.0011)·dietary α-tocopherol [mg kg^?1)]×dietary MUFA/PUFA (P<0.0067)), and hexanal concentration in meat (hexanal [ng·g^?1]=14807.9 (±1489.8)?28.8 (±10.6) dietary α-tocopherol [mg·kg^?1] (P<0.01)?8436.6 (±1701.6)·dietary MUFA/PUFA (P<0.001)+24.0 (±11.22)·dietary α-tocopherol·dietary MUFA/PUFA (P<0.0416)). It is concluded that partial substitution of dietary PUFA with MUFA lead to an increase in the concentration of α-tocopherol in muscle and microsome extracts. An interaction between dietary α-tocopherol and fatty acids exists, in which at low level of dietary vitamin E inclusion, a low MUFA/PUFA ratio leads to a reduction in the concentration of α-tocopherol in microsome extracts and a concentration of hexanal in meat above the expected values.     

Quirks of dye nomenclature. 5. Rhodamines

Rhodamines were first produced in the late 19^th century, when they constituted a new class of synthetic dyes. These compounds since have been used to color many things including cosmetics, inks, textiles, and in some countries, food products. Certain rhodamine dyes also have been used to stain biological specimens and currently are widely used as fluorescent probes for mitochondria in living cells. The early history and current biological applications are sketched briefly and an account of the ambiguities, complications and confusions concerning dye identification and nomenclature are discussed.     

Quirks of dye nomenclature. 6. Malachite green

Malachite green was discovered independently by two researchers in Germany in the 19^th century and found immediate employment as a dye and a pigment. Subsequently, other uses, such as staining biological specimens, emerged. A much later application was the control of fungal and protozoan infections in fish, for which the dye remains popular, although illegal in many countries owing to a variety of toxicity problems. In solution, malachite green can exist as five different species depending on the pH. The location of the positive charge of the colored cation on a carbon atom or a nitrogen atom is still debated. The original names of this dye, and their origins, are briefly surveyed.