Recruitment of tumor necrosis factor receptor-associated factor family proteins to apoptosis signal-regulating kinase 1 signalosome is essential for oxidative stress-induced cell death

Apoptosis signal-regulating kinase 1 (ASK1) plays a pivotal role in oxidative stress-induced cell death. Reactive oxygen species disrupt the interaction of ASK1 with its cellular inhibitor thioredoxin and thereby activates ASK1. However, the precise mechanism by which ASK1 freed from thioredoxin undergoes oligomerization-dependent activation has not been fully elucidated. Here we show that endogenous ASK1 constitutively forms a high molecular mass complex including Trx ( approximately 1,500-2,000 kDa), which we designate ASK1 signalosome. Upon H(2)O(2) treatment, the ASK1 signalosome forms a higher molecular mass complex at least in part because of the recruitment of tumor necrosis factor receptor-associated factor 2 (TRAF2) and TRAF6. Consistent with our previous findings that TRAF2 and TRAF6 activate ASK1, H(2)O(2)-induced ASK1 activation and cell death were strongly reduced in the cells derived from Traf2-/- and Traf6-/- mice. A novel signaling complex including TRAF2, TRAF6, and ASK1 may thus be the key component in oxidative stress-induced cell death.     

Microbial diversity of soils on the banks of the Solimões and Negro rivers, state of Amazonas, Brazil

Analysis of bacterial diversity in soils along the banks of the Solimões and Negro rivers, state of Amazonas, Brazil, was by partial sequencing of the genes codifying the rDNA16S region. Diversity of operational taxonomic units (OTU) and of the divergent sequences obtained were applied in comparative analysis of microbiological diversity in the two environments, based on richness estimators and OTU diversity indices. The higher OTU diversity in the Solimões was based on the higher number of parameters that evoke this. The interaction between the nucleotide sequences of bacteria inhabiting the two riverine environments indicated that the two microrganism communities are similar in composition.     

Generation of knock-in mice carrying third cones with spectral sensitivity different from S and L cones

Red-green color vision in primates is unique in the sense that it is mediated by two photoreceptor cells that are indistinguishable in all aspects except for their visual pigments. In order to generate an animal model for investigation of the interaction between red-green inputs at the molecular level, we applied knock-in technology and X-chromosome inactivation machinery to make a mouse model with cone cells possessing visual pigments with different spectral sensitivities. We introduced a S308A point mutation into the Green opsin gene allele on the X-chromosome. This manipulation generated a 24 nm red-shift of absorption maximum in the cone pigment with negligible functional differences in other molecular properties. Amplitudes of responses in ERG and ganglion cell recordings of homozygotes were similar to those of wild-types, although the spectral sensitivities differed. Heterozygotes showed variable spectral sensitivities of ganglion cell responses due to the different integration of the native and the S308A cone inputs on the dendritic fields. In situ hybridization experiments showed that cone cells with respective pigments formed patch-like clusters of specific L cone-types, approximately 30 mum in diameter, which were randomly distributed in the dorsal region of the retinas. Since the patch-like clustering was arranged by X-inactivation, such clustering could be present in the peripheral retinas of New World monkeys with polymorphic L pigments, indicating that our mice would be a suitable model to study evolution of the mammalian color vision system.     

Common antigenic determinant in the receptor binding sites of Escherichia coli enterotoxin and cholera toxin

Abstract A mutant (TUH No. 9) of a porcine strain of enterotoxigenic Escherichia coli (ETEC) produces as abnormal B subunit (B') of heat-labile enterotoxin (LT), which has aspartate instead of glycine at residue 33 from the N-terminus and does not bind to the receptor, GM1 ganglioside. The antigenicities of the receptor-binding site of LT were analyzed.
The antibody, which could not bind to the B' subunit in the anti-B subunit of porcine LT(LTp)-serum, could bind to cholera toxin (CT), LTp and LT produced by a human ETEC strain (LTh), suggesting that it recognizes a common epitope of LTp, LTh and CT. Thus glycine at residue 33 from the N-terminus in the B subunit of CT, LTh and LTp may be related to the common epitope of these three toxins. The bindings of CT, LTh and LTp to the antibody were inhibited by the GM1 ganglioside.
These data indicate that the antibody recognizes a common epitope in the receptor (GM1 ganglioside)-binding site of CT, LTh and LTp.     

Singlet oxygen quenching activity of human serum

Singlet oxygen is regarded as contributing to the pathogenesis of various diseases including light-induced skin disorders and inflammatory response. In this study, the correlation between singlet oxygen quenching activity (SOQA) of human serum and blood biochemistry or life-style was evaluated. Healthy volunteers were recruited and carried out a measurement of SOQA by using electron paramagnetic resonance (EPR) and a questionnaire survey about a smoking. It was demonstrated that major quenchers of singlet oxygen in serum are proteins, and small molecular anti-oxidants relatively play a minor role. SOQA of whole sera showed no correlation with protein concentration, but positively correlated with SOQA of small molecular fraction. In vitro studies demonstrated that the decrease of sulfhydryl groups by NO or superoxide significantly attenuated SOQA of albumin. Together, these results may imply that the underlying oxidative condition in each individual influences both small molecular antioxidant states and the sulfhydryl content of serum proteins. SOQA of sera from women with a smoking history was significantly lower compared to non-smoking women, suggesting that the smoking habit impaired the defense mechanism against singlet oxygen.     

l-Pantoyl lactone dehydrogenase from Rhodococcus erythropolis: genetic analyses and application to the stereospecific oxidation of l-pantoyl lactone

The 1,2-propanediol (1,2-PD) inducible membrane-bound L-pantoyl lactone (L-PL) dehydrogenase (LPLDH) has been isolated from Rhodococcus erythropolis AKU2103 (Kataoka et al. in Eur J Biochem 204:799, 1992). Based on the N-terminal amino acid sequence of LPLDH and the highly conserved amino acid sequence in homology search results, the LPLDH gene (lpldh) was cloned. The gene consists of 1,179 bases and encodes a protein of 392 amino acid residues. The deduced amino acid sequence showed high similarity to the proteins of the FMN-dependent α-hydroxy acid dehydrogenase/oxidase family. The overexpression vector pKLPLDH containing lpldh with its upstream region (1,940 bp) was constructed and introduced into R. erythropolis AKU2103. The recombinant R. erythropolis AKU2103 harboring pKLPLDH showed six times higher LPLDH activity than the wild-type strain. Conversion of L-PL to ketopantoyl lactone was achieved with 92% or 80% conversion yield when the substrate concentration was 0.768 or 1.15 M, respectively. Stereoinversion of L-PL to D-PL was also carried out by using the combination of recombinant R. erythropolis AKU2103 harboring pKLPLDH and ketopantoic acid-reducing Escherichia coli.     

Movement patterns of adult Sakhalin taimen,Parahucho perryi,between stream habitats of the Bekanbeushi River system,eastern Hokkaido,Japan

The behavior of endangered adult Sakhalin taimen Parahucho perryi was tracked during 2008–2010 using acoustic telemetry in the Bekanbeushi River system, which flows through eastern Hokkaido, Japan. Movement distances per unit time of tracked P. perryi were compared between mid- and downstream habitats. Results indicated that movement distances were significantly longer in downstream habitats during all seasons. The movement distances in each stream decreased from spring to autumn. Moreover, tracked P. perryi exhibited crepuscular movement patterns; however, patterns were less pronounced in downstream habitats than in up- and midstream habitats. These findings strongly suggested that adult P. perryi exhibit nearly distinctive movement patterns across stream habitats; thus, fish may adopt different foraging tactics in each stream habitat. Fish moved more frequently in spring, which included the post-wintering and post-spawning season, most likely to search for food and more desirable habitat for recovery.     

Identification of a novel galactosyl transferase involved in biosynthesis of the mycobacterial cell wall

The possibility of the Rv3782 protein of Mycobacterium tuberculosis being a putative galactosyl transferase (GalTr) implicated in galactan synthesis arose from its similarity to the known GalTr Rv3808c, its classification as a nucleotide sugar-requiring inverting glycosyltransferase (GT-2 family), and its location within the "possible arabinogalactan biosynthetic gene cluster" of M. tuberculosis. In order to study the function of the enzyme, active membrane and cell wall fractions from Mycobacterium smegmatis containing the overexpressed Rv3782 protein were incubated with endogenous decaprenyldiphosphoryl-N-acetylglucosaminyl-rhamnose (C(50)-P-P-GlcNAc-Rha) as the primary substrate for galactan synthesis and UDP-[(14)C]galactopyranose as the immediate precursor of UDP-[(14)C]galactofuranose, the ultimate source of all of the galactofuranose (Galf) units of galactan. Obvious increased and selective synthesis of C(50)-P-P-GlcNAc-Rha-Galf-Galf, the earliest product in the pathway leading to the fully polymerized galactan, was observed, suggesting that Rv3782 encodes a GalTr involved in the first stages of galactan synthesis. Time course experiments pointed to a possible bifunctional enzyme responsible for the initial synthesis of C(50)-P-P-GlcNAc-Rha-Galf, followed by immediate conversion to C(50)-P-P-GlcNAc-Rha-Galf-Galf. Thus, Rv3782 appears to be the initiator of galactan synthesis, while Rv3808c continues with the subsequent polymerization events.     

Impairment of spatial learning by estradiol treatment in female mice is attenuated by estradiol exposure during development

High doses of estradiol (E(2)) can impair spatial learning in the Morris water maze, in ovariectomized mice, but the same dose has no effect on adult castrated males. Here, we test the hypothesis that this sex difference is caused by neonatal actions of E(2). In Experiment 1, C57BL/6J pups were given daily estradiol benzoate (EB) or oil injections from the day of birth until postnatal Day 3. Adults were gonadectomized and received EB (s.c.) or oil 28 h before the first day of training, and 4 h before each of four daily training sessions on the Morris water maze. Females given oil as neonates, and EB prior to training displayed the poorest performance. Females that received EB as neonates and EB prior to training were insensitive to the deleterious effects of adult EB and performed better than males given the same hormone treatments. We conducted a second experiment using aromatase enzyme knockout (ArKO) mice. Adult male and female ArKO and wild-type (WT) littermates were gonadectomized and received either injections of oil or EB prior to and during water maze training (as described above). Hormone treatment failed to affect performance, yet, female but not male ArKO mice showed impaired learning compared to WT littermates. Thus, exposure to estradiol during neonatal development can counteract the deleterious effects of EB on adult spatial learning.