Growth factor-dependent branching of the ureteric bud is modulated by selective 6-O sulfation of heparan sulfate

Heparan sulfate proteoglycans (HSPGs) are found in the basement membrane and at the cell-surface where they modulate the binding and activity of a variety of growth factors and other molecules. Most of the functions of HSPGs are mediated by the variable sulfated glycosaminoglycan (GAG) chains attached to a core protein. Sulfation of the GAG chain is key as evidenced by the renal agenesis phenotype in mice deficient in the HS biosynthetic enzyme, heparan sulfate 2-O sulfotransferase (Hs2st; an enzyme which catalyzes the 2-O-sulfation of uronic acids in heparan sulfate). We have recently demonstrated that this phenotype is likely due to a defect in induction of the metanephric mesenchyme (MM), which along with the ureteric bud (UB), is responsible for the mutually inductive interactions in the developing kidney (Shah et al., 2010). Here, we sought to elucidate the role of variable HS sulfation in UB branching morphogenesis, particularly the role of 6-O sulfation. Endogenous HS was localized along the length of the UB suggesting a role in limiting growth factors and other molecules to specific regions of the UB. Treatment of cultures of whole embryonic kidney with variably desulfated heparin compounds indicated a requirement of 6O-sulfation in the growth and branching of the UB. In support of this notion, branching morphogenesis of the isolated UB was found to be more sensitive to the HS 6-O sulfation modification when compared to the 2-O sulfation modification. In addition, a variety of known UB branching morphogens (i.e., pleiotrophin, heregulin, FGF1 and GDNF) were found to have a higher affinity for 6-O sulfated heparin providing additional support for the notion that this HS modification is important for robust UB branching morphogenesis. Taken together with earlier studies, these findings suggest a general mechanism for spatio-temporal HS regulation of growth factor activity along the branching UB and in the developing MM and support the view that specific growth factor-HSPG interactions establish morphogen gradients and function as developmental switches during the stages of epithelial organogenesis (Shah et al., 2004).     

Characterization and potential application of purified aldehyde oxidase from Pseudomonas stutzeri IFO12695

The molecular weight of purified aldehyde oxidase from Pseudomonas stutzeri IFO12695 was estimated to be 160 kDa by a gel filtration method. SDS-PAGE showed that the enzyme consisted of three non-identical subunits with molecular weights of 18, 38, and 83 kDa. The enzyme exhibited an absorption spectrum with maxima at 277, 325, 365, 415, 450, 480, and 550 nm and possessed molybdenum, CMP, iron, sulfur, and FAD as its cofactors, indicating that it belonged to the xanthine oxidase family. A variety of aliphatic and aromatic aldehydes were oxidized; and among them n-hexylaldehyde gave the most rapidly action. When 10 mM formaldehyde was treated with the aldehyde oxidase in the presence of catalase for 240 min, the formaldehyde concentration was reduced to 0.8 mM, suggesting this enzyme might be effective for the removal of formaldehyde contained in wastewater.     

Lineage admixture during postglacial range expansion is responsible for the increased gene diversity of Kalopanax septemlobus in a recently colonised territory

We aimed to reveal the effects of range expansion and subsequent lineage admixture from separated glacial refugia on genetic diversity of Kalopanax septemlobus in Japan, by combining nuclear microsatellite data and ecological niche modelling. Allelic richness and gene diversity were compared at the population and regional level. We also statistically examined these indices as a function of population accessibility to the last glacial maximum (LGM) palaeodistribution reconstructed by ecological niche modelling to test a simple range expansion scenario from glacial refugia. Genetic diversity was highest in the populations of southern Japan and gradually decreased towards the north. However, an additional centre of genetic diversity, when measured as gene diversity, was found in northern Honshu Island, where distinct lineages were shown to be in contact. Positive effects of population accessibility to the LGM range were detected in both diversity indices at different spatial scales. The combined data support independent postglacial range expansions towards the north from the edge populations on the exposed coastal shelf of Pacific and Sea of Japan in northern Honshu during the LGM, which subsequently resulted in markedly low genetic diversity in the northernmost extant range, Hokkaido. The regional increase in gene diversity in northern Honshu is likely to be the result of postglacial lineage admixture. Relative difference in the spatial scales best relating population genetic diversity with the LGM distribution can be explained by a higher rate of allelic richness diversity loss during range expansions and stronger effects of lineage admixture on gene diversity.     

Distribution pattern of Polar cod (<Emphasis Type="Italic">Boreogadus saida</Emphasis>) larvae and larval fish assemblages in relation to oceanographic parameters in the northern Bering Sea and Chukchi Sea

Polar cod, Boreogadus saida, is a key species in the Arctic Ocean ecosystem. We examined the distribution pattern of B. saida and other fish larvae in relation to oceanographic parameters, including sea surface temperature and salinity (SST and SSS), the mode of temperature and salinity within the water column (F _temp and F _sal), and the temporal duration between the date of sea ice retreat and the date of field surveys (dSRT) in the northern Bering Sea and Chukchi Sea during the summers of 2008 and 2013. Sampling was conducted onboard the T/S Oshoro-Maru using a bongo net for 2 years. At sampling stations, the temperature and salinity were measured using conductivity-temperature-depth profiler casts. We calculated dSRT from satellite derived polar gridded sea ice concentration data. A total of 1186 individuals comprising 7 families and 16 species were collected, with B. saida (35 %) and Ammodytes hexapterus (27 %) dominating the catch in number. Based on the species composition (cluster analysis), the sampling stations were divided into four groups. Pleuronectidae dominated group A, which was characterized by relatively high temperature (SST and F _temp), while B. saida dominated group B, characterized by low temperatures. A. hexapterus dominated group C and D, which had similar temperatures and salinities, but group C had a shorter dSRT than group D. The latter was also differentiated by the presence of Lumpenus sp. B. saida, which were most abundant in regions where temperatures ranged from ?2 to 0.5 °C. These results, the geographical variation of group B sampling stations, suggest there are two separate spawning areas characterized by similar fish communities and related to consistently cold bottom temperatures. Moreover, stations with a higher abundance of smaller sized larval B. saida were characterized by a short dSRT, whereas stations with a lower abundance and a larger size were characterized by a long dSRT.     

Relationship between susceptibility and immune response to staphylococcal exfoliative toxin A in mammalian species

Staphylococcal exfoliative toxin A (ETA) had a splitting effect at the granular layer of skin in humans and neonatal mice, but not in rabbits, guinea pigs, golden hamsters, or rats. Besides its splitting effect, ETA could stimulate productions of neutralizing antibody to ETA in rabbits, rats and B10D2 mice, but not in golden hamsters, guinea pigs, or ICR, HRS/J, and C57BL/10 mice. In our epidemiological investigation of human sera, the percentage of antibody to ETA in sera obtained from patients with impetigo (8%) was lower than those in sera of healthy males (23%) and females (29%). The relationship between susceptibility and immune response to ETA in these mammalians could be divided into three groups: the possession of resistant skin and high production of antibody to ETA; the possession of resistant skin and low production of antibody to ETA; the possession of sensitive skin and various titers of antibody to ETA.     

Non-enzymatic glycation of antithrombin III in vitro

Non-enzymatic glycation of antithrombin III (AT-III) has been proposed as a significant contributor to the increased incidence of thrombo-occlusive events in diabetics. AT-III, isolated from normal human plasma by means of heparin affinity and ion-exchange chromatography, was incubated with 0-0.5 M glucose in neutral phosphate buffer at 37 degrees C. The extent of non-enzymatic glycation could be monitored by uptake of radioactivity as well as by binding to a phenylboronate affinity resin, which effectively retards AT-III containing ketoamine-linked glucose. Non-enzymatically glycated AT-III (approx. 1 mol glucose/mol protein) bound heparin nearly as efficiently as non-glycated AT-III. The two AT-III preparations were equally active in inhibiting thrombin cleavage of chromogenic substrate. Following incubation with [14C]glucose, structural analyses of cyanogen-bromide-cleaved peptides of enzymatically glycated AT-III showed that the [14C]glucose adducts were distributed over many sites on the molecule. This lack of specificity contrasts with the restricted sites of modification on hemoglobin, albumin and ribonuclease A, and explains why non-enzymatic glycation of AT-III has little if any effect on its function.     

Prophage Origin of a Virulent Phage Appearing on Fermentations of Lactobacillus casei S-1

For protection from the abnormal fermentation of Lactobacillus casei S-1 caused by contamination of a virulent phage, FSV, the origin of this phage was studied. Morphologies, viral structural proteins, and DNA structures of three independent isolates of FSV were compared with those of FSW, which is lysogenized in strain S-1. The results showed (i) that the morphology of FSV phages is indistinguishable from that of FSW and (ii) that all viral structural components found in FSW are present in the particles of FSV's. In addition, restriction endonuclease analyses of viral DNA showed that the HindIII-digested fragments of FSW DNA, the sum of which covered at least 94.7% of this phage genome, were conserved in the FSV DNA digests. Results of Southern filter hybridization of the S-1 and prophage-cured cell (C239) DNAs with FSV DNA as a probe revealed that C239 had lost most of the FSV DNA sequence, whereas S-1 had about one copy of the FSV DNA sequence. These results indicate that virulent phage FSV is derived from the lysogenized phage FSW. Therefore, the appearance of FSV can be eliminated by using the prophage-cured derivative of S-1.     

Evolution of restriction sites of ribosomal DNA in natural populations of the field mouse,Apodemus speciosus

An analysis by restriction endonuclease digestion of ribosomal DNA (rDNA) was carried out in natural populations of Apodemus speciosus, a field mouse that is endemic to Japan. Two restriction sites, the EcoRI (E3) and DraI (D4) sites, in the nontranscribed spacer region downstream from the gene for 28S RNA showed polymorphism within and between individuals in the populations from the Japanese main islands. By contrast, populations from the small adjoining islands which are thought to have separated from the main islands 1–2 × 10⁴ years ago showed relatively low levels of polymorphism within and between individuals, i.e., one of the polymorphic bands in the case of each enzyme was predominant in these populations, irrespective of the variants. These results indicate that the rate of fixation of site variations depends on population size and that the direction of fixation is random. Furthermore, each polymorphic restriction site seems to be fixed independently.Correspondence to: H. Suzuki     

Distributions of cadmium,zinc, and polyphenols in Gamblea innovans

Gamblea innovans is a Cd- and Zn-accumulating deciduous tree widely distributed in the secondary forests of Japan. We aimed to understand the characteristics of Cd and Zn accumulation in G. innovans in order to effectively utilize the species for phytoremediation. To accomplish that, we studied the relationship between secondary metabolite concentrations and the accumulation and distributions of Cd and Zn in G. innovans leaves and basal stems using micro-X ray fluorescence (µ-XRF). Our results showed a negative correlation between Zn leaf concentrations and polyphenol/2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity. This finding might be related to stress or the manifestation of a mechanism for tolerance to Cd and Zn accumulation. In addition, we observed that Cd accumulated primarily in the apoplastic region of surface tissues such as bark and the epidermis of leaves, whereas Zn accumulated in both apoplastic and symplastic regions. Thus, it might be possible that G. innovans can distinguish between Cd and Zn and control their translocation.     

Hydration of oligosaccharides: anomalous hydration ability of trehalose.

The disaccharide trehalose extensively exists in anhydrobiotic organism and is considered to play an important role in preserving the integrity of biomembrane. However, the preserving mechanism remains unclear. In this report, we examine the hydration abilities of trehalose and several oligosaccharides composed of alpha-D-glucopyranosyl residues. The unfrozen water fraction per molecule was determined from differential scanning calorimetry measurements of their aqueous solutions. Further, the NMR relaxation time of the natural abundance 17O of water is measured for several saccharide solutions. These results indicate that trehalose has the highest hydration ability among the saccharides studied. In other words, trehalose can effectively lower the mobility of water molecules hydrogen-bonded with saccharides. It is thus reasonable that, among the disaccharides studied, trehalose exhibits the maximum stabilizing effect on the bilayer structure of lipid whose acyl chains are bonded with each other by the apolar interaction, because the apolar interaction is strengthened with the stabilization of the surrounding water structure.