Polyphenol‐solubility alters amyloid fibril formation of α‐synuclein

Amyloid fibril formation is associated with various amyloidoses, including neurodegenerative diseases such as Alzheimer''s and Parkinson''s diseases. Amyloid fibrils form above the solubility of amyloidogenic proteins or peptides upon breaking supersaturation, followed by a nucleation and elongation mechanism, which is similar to the crystallization of solutes. Many additives, including salts, detergents, and natural compounds, promote or inhibit amyloid formation. However, the underlying mechanisms of the opposing effects are unclear. We examined the effects of two polyphenols, that is, epigallocatechin gallate (EGCG) and kaempferol‐7─O─glycoside (KG), with high and low solubilities, respectively, on the amyloid formation of α‐synuclein (αSN). EGCG and KG inhibited and promoted amyloid formation of αSN, respectively, when monitored by thioflavin T (ThT) fluorescence or transmission electron microscopy (TEM). Nuclear magnetic resonance (NMR) analysis revealed that, although interactions of αSN with soluble EGCG increased the solubility of αSN, thus inhibiting amyloid formation, interactions of αSN with insoluble KG reduced the solubility of αSN, thereby promoting amyloid formation. Our study suggests that opposing effects of polyphenols on amyloid formation of proteins and peptides can be interpreted based on the solubility of polyphenols.     

Whole-body to tissue concentration ratios for use in biota dose assessments for animals

Environmental monitoring programs often measure contaminant concentrations in animal tissues consumed by humans (e.g., muscle). By comparison, demonstration of the protection of biota from the potential effects of radionuclides involves a comparison of whole-body doses to radiological dose benchmarks. Consequently, methods for deriving whole-body concentration ratios based on tissue-specific data are required to make best use of the available information. This paper provides a series of look-up tables with whole-body:tissue-specific concentration ratios for non-human biota. Focus was placed on relatively broad animal categories (including molluscs, crustaceans, freshwater fishes, marine fishes, amphibians, reptiles, birds and mammals) and commonly measured tissues (specifically, bone, muscle, liver and kidney). Depending upon organism, whole-body to tissue concentration ratios were derived for between 12 and 47 elements. The whole-body to tissue concentration ratios can be used to estimate whole-body concentrations from tissue-specific measurements. However, we recommend that any given whole-body to tissue concentration ratio should not be used if the value falls between 0.75 and 1.5. Instead, a value of one should be assumed.     

Purification and characterization of the plasmodial phosphatase that hydrolyses the phosphorylated light chain of Physarum myosin II from Physarum polycephalum

A phosphatase was purified through a combination of ion‐exchange and hydrophobic chromatography followed by native PAGE from Physarum plasmodia. Recently, we demonstrated that this phosphatase isoform has a hydrolytic activity towards the PMLC (phosphorylated light chain of Physarum myosin II) at pH 7.6. The apparent molecular mass of the purified enzyme was estimated at approximately 50 kDa by means of analytical gel filtration. The enzyme was purified 340‐fold to a final phosphatase activity of 400 pkat/mg of protein. Among the phosphorylated compounds tested for hydrolytic activity at pH 7.6, the enzyme showed no activity towards nucleotides. At pH 7.6, hydrolytic activity of the enzyme against PMLC was detected; at pH 5.0, however, no hydrolytic activity towards PMLC was observed. The K _m of the enzyme for PMLC was 10 μM, and the V _max was 1.17 nkat/mg of protein. Ca²⁺ (10 μM) inhibited the activity of the enzyme, and Mg²⁺ (8.5 μM) activated the dephosphorylation of PMLC. Mn²⁺ (1.6 μM) highly stimulated the enzyme's activity. Based on these results, we concluded that the enzyme is likely to be a phosphatase with hydrolytic activity towards PMLC.     

A bacterial elongation factor G homologue exclusively functions in ribosome recycling in the spirochaete Borrelia burgdorferi

Translation elongation factor G (EF‐G) in bacteria plays two distinct roles in different phases of the translation system. EF‐G catalyses the translocation of tRNAs on the ribosome in the elongation step, as well as the dissociation of the post‐termination state ribosome into two subunits in the recycling step. In contrast to this conventional view, it has very recently been demonstrated that the dual functions of bacterial EF‐G are distributed over two different EF‐G paralogues in human mitochondria. In the present study, we show that the same division of roles of EF‐G is also found in bacteria. Two EF‐G paralogues are found in the spirochaete Borrelia burgdorferi, EF‐G1 and EF‐G2. We demonstrate that EF‐G1 is a translocase, while EF‐G2 is an exclusive recycling factor. We further demonstrate that B. burgdorferi EF‐G2 does not require GTP hydrolysis for ribosome disassembly, provided that translation initiation factor 3 (IF‐3) is present in the reaction. These results indicate that two B. burgdorferi EF‐G paralogues are close relatives to mitochondrial EF‐G paralogues rather than the conventional bacterial EF‐G, in both their phylogenetic and biochemical features.     

Comparative genomic analysis of 1047 completely sequenced cDNAs from an Arabidopsis-related model halophyte, <Emphasis Type="Italic">Thellungiella halophila</Emphasis>

Background  

Thellungiella halophila (also known as T. salsuginea) is a model halophyte with a small size, short life cycle, and small genome. Thellungiella genes exhibit a high degree of sequence identity with Arabidopsis genes (90% at the cDNA level). We previously generated a full-length enriched cDNA library of T. halophila from various tissues and from whole plants treated with salinity, chilling, freezing stress, or ABA. We determined the DNA sequences of 20 000 cDNAs at both the 5'- and 3' ends, and identified 9569 distinct genes.     

Development of a novel fluorometric assay for nitric oxide utilizing sesamol and its application to analysis of nitric oxide‐releasing drugs

Nitric oxide (NO) is related to various physiological effects as well as to numerous diseases caused by accentuation of NO production. Measurement of NO in cells and tissues is difficult as NO readily reacts with other molecules; furthermore, its half‐life as a radical is fleeting. Currently, many NO pharmaceuticals are marketed as therapeutic agents for ischemic disease. Consequently, the identification of NO radicals and determination of generation rate from pharmaceuticals is very important when the effect of the medicinal supply is estimated. In this study, we developed a fluorometric assay for NO employing sesamol (3,4‐methylenedioxyphenol) as a fluorometric substrate. Sesamol is converted to a fluorescent derivative (ex. 365 nm, em. 447 nm), which is dimmer in the presence of NO. The detection limit of NO with this method is 400 fmol; moreover, NO generated from drugs can be measured. Copyright © 2009 John Wiley & Sons, Ltd.     

The effect of photosynthetically active radiation and temperature on the photosynthesis of two Vietnamese species of Sargassum,S. mcclurei and S. oligocystum,based on the field and laboratory measurements

SUMMARY The effects of photosynthetically active radiation (PAR) and temperature on the photosynthesis of two Vietnamese brown algae, Sargassum mcclurei and S. oligocystum (Fucales), were determined by field and laboratory measurements. Dissolved oxygen sensors and pulse‐amplitude modulated (PAM) fluorometry were used for the measurements of photosynthetic efficiency. A Diving‐PAM revealed that underwater measurements of the effective quantum yield (Φ _PSII ) of both species declined with increasing incident PAR, with minimum Φ _PSII occurring during noon to early afternoon. Φ _PSII recovered in the evening, indicating photo‐adaptation to excessive PAR. In laboratory experiments, Φ _PSII also decreased under continuous exposure to 1000 μmol photons m^?2 s^?1; and full recovery occurred after 12 h of dark acclimatization. The net photosynthesis – PAR experiments of S. mcclurei and S. oligocystum conducted at 28°C revealed that the net photosynthetic rate quickly increased at PAR below the saturation irradiance of 361 and 301 μmol photons m^?2 s^?1 and nearly saturated to maximum net photosynthetic rates of 385 and 292 μg O₂ g_ww ^{? 1} min^?1 without photoinhibition, respectively. Gross photosynthesis and dark respiration experiments determined over a range of temperatures (12–40°C), revealed that the maximum gross photosynthetic rates of 201 and 147 μg O₂ g_ww ^{? 1} min^?1 occurred at 32.9 and 30.7°C for S. mcclurei and S. oligocystum, respectively. The dark respiration rates increased exponentially over the temperature ranges examined. The estimated maximum value of the maximum quantum yield occurred at 19.3 and 20.0°C and was 0.76 and 0.74, respectively. Similar to the natural habitat of the study site, these two species tolerated the relatively high temperatures and broad range of PAR. The ability of these species to recover from exposure to high PAR is one of the mechanisms that allow them to flourish in the shallow water environment.     

Defective immune responses in mice lacking LUBAC‐mediated linear ubiquitination in B cells

The linear ubiquitin chain assembly complex (LUBAC) plays a crucial role in activating the canonical NF‐κB pathway, which is important for B‐cell development and function. Here, we describe a mouse model (B‐HOIP^Δlinear) in which the linear polyubiquitination activity of LUBAC is specifically ablated in B cells. Canonical NF‐κB and ERK activation, mediated by the tumour necrosis factor (TNF) receptor superfamily receptors CD40 and TACI, was impaired in B cells from B‐HOIP^Δlinear mice due to defective activation of the IKK complex; however, B‐cell receptor (BCR)‐mediated activation of the NF‐κB and ERK pathways was unaffected. B‐HOIP^Δlinear mice show impaired B1‐cell development and defective antibody responses to thymus‐dependent and thymus‐independent II antigens. Taken together, these data suggest that LUBAC‐mediated linear polyubiquitination is essential for B‐cell development and activation, possibly via canonical NF‐κB and ERK activation induced by the TNF receptor superfamily, but not by the BCR.     

Platelets Promote Tumor Growth and Metastasis via Direct Interaction between Aggrus/Podoplanin and CLEC-2

The platelet aggregation-inducing factor Aggrus, also known as podoplanin, is frequently upregulated in several types of tumors and enhances hematogenous metastasis by interacting with and activating the platelet receptor CLEC-2. Thus, Aggrus–CLEC-2 binding could be a therapeutic molecular mechanism for cancer therapy. We generated a new anti-human Aggrus monoclonal antibody, MS-1, that suppressed Aggrus–CLEC-2 binding, Aggrus-induced platelet aggregation, and Aggrus-mediated tumor metastasis. Interestingly, the MS-1 monoclonal antibody attenuated the growth of Aggrus-positive tumors in vivo. Moreover, the humanized chimeric MS-1 antibody, ChMS-1, also exhibited strong antitumor activity against Aggrus-positive lung squamous cell carcinoma xenografted into NOD-SCID mice compromising antibody-dependent cellular cytotoxic and complement-dependent cytotoxic activities. Because Aggrus knockdown suppressed platelet-induced proliferation in vitro and tumor growth of the lung squamous cell carcinoma in vivo, Aggrus may be involved in not only tumor metastasis but also tumor growth by promoting platelet-tumor interaction, platelet activation, and secretion of platelet-derived factors in vivo. Our results indicate that molecular target drugs inhibiting specific platelet–tumor interactions can be developed as antitumor drugs that suppress both metastasis and proliferation of tumors such as lung squamous cell carcinoma.