Repetitive elements in the major histocompatibility complex (MHC) class I region of a teleost, medaka: identification of novel transposable elements

The repetitive elements of medaka (Oryzias latipes) are poorly characterized in spite of recent rapid progress in the medaka genome analysis. Here we report the characterization of the repetitive elements in the major histocompatibility complex (MHC) class I region, which spans about 400 kb and is one of the best characterized regions of the medaka genome. Microsatellite, low complexity regions, transposable elements, and other repeats occupied 0.68, 0.98, 7.0 and 2.9%, respectively, of the MHC class I region. Eleven transposable elements, three LTR-type, six LINE-type and two DNA-type, including several novel ones, were identified. Genomic Southern hybridization analysis indicated that these LINE-type and DNA-type elements have many copies in the medaka genome, whereas the LTR-type elements have only several copies. The comparison of the medaka MHC class I region with those of zebrafish and fugu shows the presence of three medaka lineage-specific tandem duplications of the PSMB (proteasome beta-type subunit) 8 and class Ia genes. Since eight of the 11 transposable elements were located in this region, these elements may have played a role in the medaka-specific DNA rearrangement.     

Role of p38 mitogen-activated protein kinase in thrombus formation

The present study was designed to elucidate the role of p38 mitogen-activated protein kinase (p38) in thrombus formation. We used p38alpha heterozygous (p38alpha+/-) mice and used ferric chloride (FeCl3)-induced carotid artery injury as a model of thrombus formation. The time to thrombotic occlusion induced by FeCl3 in p38alpha+/- mice was prolonged compared to that in wild-type (WT) mice. Platelets prepared from p38alpha+/- mice showed impairment of the aggregatory response to a low concentration of U46619, a thromboxane A2 analogue. Furthermore, platelets prepared from p38alpha+/- mice and activated by U46619 were poorly bound to fibrinogen compared with those from WT mice. Both the expression and activity of tissue factor induced by FeCl3 in WT mice were higher than those in p38alpha+/- mice. These results suggest that p38 plays an important role in thrombus formation by regulating platelet function and tissue factor activity.     

Rheological properties and molecular structure of tunicate cellulose in LiCl/1,3-dimethyl-2-imidazolidinone

Solution properties and molecular structure of tunicate cellulose (TC), an animal cellulose from Halocynthia roretzi, were investigated in terms of rheological and dilute solution properties. The solvent used is 8 wt % LiCl/1,3-dimethyl-2-imidazolidinone (DMI). A solution of dissolving pulp (DP), derived from plant, was also used for comparison. The weight-average molecular weight, Mw, and the limiting viscosity number, [eta], of the TC were evaluated to be 413 x 10(6) and 2645 mL/g, respectively. The TC solution showed the same concentration dependence of GN (GN=5.49 x 10(6)phiw(2.1)4 Pa; phiw: weight fraction of cellulose in solution; GN: plateau modulus) as the DP solution and, moreover, also as the solution of cotton linter (CC) in 8 wt % LiCl/N,N-dimethylacetamide (DMAc). This exponent of 2.1(4) indicates that network structure by entanglements was formed in these solutions. According to the theory of Fetters et al., moreover, such identity means that all of these celluloses have the identical chain structure though their biological origins are far different. On the other hand, the phiw-dependence of eta0-etas (eta0=zero shear rate viscosity of solution; etas=solvent viscosity) was different between the TC and the DP solution in the semidilute regime: the TC solution exhibited eta0-etas proportional, variant phiw(7.5) and the DP solution eta0-etas proportional, variant phiw4. According to the theory of Doi-Edwards, this exponent of 4 (the DP solution) indicates that the DP behaves as flexible polymers in the solution. In contrast, the dependence for the TC solution seems unexplainable on the basis of molecular theories. This difference probably signifies the difference in the relaxation process or mechanism in entanglement systems.     

Acute waterborne cadmium uptake in rainbow trout is reduced by dietary calcium carbonate

The effects of elevated dietary calcium (as CaCO3) and acute waterborne Cd exposure (50 microg/l) on whole body uptake, tissue uptake, and internal distribution of newly accumulated Cd, Ca2+, and Na+ in juvenile rainbow trout were examined. Fish were fed with three diets (mg Ca2+/g food): 20 (control), 30 and 60 for 7 days before fluxes were measured with radiotracers. The highest dietary Ca2+ elevation reduced waterborne whole body Ca2+ uptake, but did not protect against inhibition of waterborne Ca2+ uptake by waterborne Cd. Both Ca2+-supplemented diets reduced newly accumulated Ca2+ in the gills in relation to the control treatment, but did not prevent the Cd-inhibiting effect against accumulation of new Ca2+ in most compartments. Fish fed with Ca2+-supplemented diets showed markedly lower rates of whole body uptake and internalization (in some tissues) of waterborne Cd, illustrating that, while dietary Ca2+ supplementation did not protect against the impact of waterborne Cd on waterborne Ca2+ uptake, it did protect against the uptake of Cd. Waterborne Cd had no effect on Na+ fluxes, total Cl-, and in most body compartments, newly accumulated Na+ and total Na+ were also not affected. Dietary supplementation with CaCO3 had the same protective effect as demonstrated by dietary supplementation with CaCl2 in an earlier study. Thus, the reduction of waterborne Cd uptake and internalization by dietary Ca2+ was specifically due to Ca2+ and not to the anion.     

Angiogenic activity of osteopontin-derived peptide SVVYGLR

Angiogenesis plays an important role in various pathological conditions as well as some physiological processes. Although a number of soluble angiogenic factors have been reported, extracellular matrix also has crucial effect on angiogenesis through interaction with endothelial cells. Since recent reports showed osteopontin had some angiogenic activity, the effect of the SVVYGLR peptide, novel binding motif in osteopontin molecule, on angiogenesis was examined in this study. Synthetic peptide SVVYGLR did not have proliferative effect on endothelial cells but adhesion and migration activity to endothelial cells. Furthermore, SVVYGLR had as potent activity for tube formation in three-dimensional collagen gel as vascular endothelial growth factor which is known to be the strongest angiogenic factor. Electron microscopical analysis showed a number of microvilli on the endothelial luminar surface and tight junction formation in the luminar intercellular border between endothelial cells, indicating SVVYGLR induced cell porarity and differentiation of endothelial cells. This small peptide might be expected to stimulate angiogenesis to improve some ischemic conditions in the future because of some advantages due to smaller molecular weight.     

Effects of mutations of ABCA1 in the first extracellular domain on subcellular trafficking and ATP binding/hydrolysis

ABCA1 mediates release of cellular cholesterol and phospholipid to form high density lipoprotein (HDL). The three different mutants in the first extracellular domain of human ABCA1 associated with Tangier disease, R587W, W590S, and Q597R, were examined for their subcellular localization and function by using ABCA1-GFP fusion protein stably expressed in HEK293 cells. ABCA1-GFP expressed in HEK293 was fully functional for apoA-I-mediated HDL assembly. Immunostaining and confocal microscopic analyses demonstrated that ABCA1-GFP was mainly localized to the plasma membrane (PM) but also substantially in intracellular compartments. All three mutant ABCA1-GFPs showed no or little apoA-I-mediated HDL assembly. R587W and Q597R were associated with impaired processing of oligosaccharide from high mannose type to complex type and failed to be localized to the PM, whereas W590S did not show such dysfunctions. Vanadate-induced nucleotide trapping was examined to elucidate the mechanism for the dysfunction in the W590S mutant. Photoaffinity labeling of W590S with 8-azido-[alpha-(32)P]ATP was stimulated by adding ortho-vanadate in the presence of Mn(2+) as much as in the presence of wild-type ABCA1. These results suggest that the defect of HDL assembly in R587W and Q597R is due to the impaired localization to the PM, whereas W590S has a functional defect other than the initial ATP binding and hydrolysis.     

Phosphatidylinositol 3-kinases in carcinoembryonic antigen-related cellular adhesion molecule-mediated internalization of Neisseria gonorrhoeae

Neisseria gonorrhoeae can be internalized by mammalian cells through interactions between bacterial opacity-associated (Opa) adhesins and members of the human carcinoembryonic antigen-related cellular adhesion molecule (CEACAM) family. We examined the role of phosphatidylinositol 3-kinases (PI3Ks) in gonococcal invasion of epithelial cell lines expressing either CEACAM1 or CEACAM3. CEACAM3-mediated internalization, but not that mediated by CEACAM1, was accompanied by localized and transient accumulation of the class I PI3K product phosphatidylinositol 3,4,5-trisphosphate at sites of bacterial engulfment. Inhibition of phosphatidylinositol 3-kinases reduced CEACAM3-mediated uptake but, paradoxically, led to an increase in intracellular survival of bacteria internalized via either CEACAM1 or CEACAM3, suggesting additional roles for PI3K products. Consistent with this finding, the class III PI3K product phosphatidylinositol 3-phosphate accumulated and persisted in the membrane of gonococcal phagosomes after internalization. Inhibition of PI3K blocked phagosomal acquisition of the late endosomal marker lysosome-associated membrane protein 2 and reduced phagosomal acidification. Inhibiting phagosomal acidification with concanamycin A also increased survival of intracellular gonococci. These results suggest two modes of action of phosphatidylinositol 3-kinases during internalization of gonococci: synthesis of phosphatidylinositol 3,4,5-trisphosphate is important for CEACAM3-mediated uptake, while phosphatidylinositol 3-phosphate is needed for phagosomal maturation and acidification, which are required for optimal bacterial killing.     

Classification of dynamics of a model of motor coordination and comparison with Parkinson's disease data

In our recent reports motor coordination of human lower limbs has been investigated during pedaling a special kind of ergometer which allows its left and right pedals to rotate independently. In particular, relative phase between left and right rotational-velocity waveforms of the pedals and their amplitude modulation have been analyzed for patients with Parkinson's disease (PD). Several patients showed peculiar interlimb coordination different from the regular anti-phase pattern of normal subjects. We have reported that these disordered patterns could be classified into four groups. Moreover, it has been demonstrated that a mathematical model could reproduce most of the disordered patterns. Such a model includes a schematization of the central pattern generator with two identical half-centers mutually coupled and two tonic control signals from higher motor centers, each of which inputs to one of the half-centers. Depending on the intensities of the tonic signals and on the differences between them, the model could generate a range of dynamics comparable to the clinically observed disordered patterns. In this paper, we explore the dynamics of the model by varying the intensities of the tonic signals in the model. Using the same method used for classifying the clinical data, the dynamics of the model are classified into several groups. The classified groups for the simulated data are compared with those for the clinical data to look at qualitative correspondence. Our systematic exploration of the model's dynamics in a wide range of the parameter space has revealed global organization of the bifurcations including Hopf bifurcations and cascades of period-doubling bifurcations among others, suggesting that the bifurcations, induced by instability of stable dynamics of the human motor control system, are responsible for the emergence of the disordered coordination in PD patients.     

Isolation and phylogenetic characterization of bacteria capable of inducing differentiation in the green alga Monostroma oxyspermum

Many green algae cannot develop normally when they are grown under axenic conditions. Monostroma oxyspermum, for example, proliferates unicellularly in an aseptic culture, but develops into a normal foliaceous gametophyte in the presence of some marine bacteria. More than 1000 bacterial strains were isolated from marine algae and sponges and assayed for their ability to induce the morphogenesis of unicellular M. oxyspermum. Fifty bacterial strains exhibiting morphogenesis-inducing activity against unicellular M. oxyspermum were isolated. The partial gyrB (approximately 1.2 kbp) and 16S rDNA (approximately 1.4 kbp) sequences of about 40 active strains were determined, and their phylogenetic relationships were analysed. All these strains were located within the Cytophaga-Flavobacterium-Bacteroides (CFB) complex, and most of these strains were clustered in a clade comprising Zobellia uliginosa. On the other hand, these bacteria also exhibited morphogenetic activity against germ-free spores of Ulva pertusa, Ulva conglobata and Enteromorpha intestinalis. Moreover, these bacteria induced the release of spores from the leafy young gametophyte of M. oxyspermum. These results indicate that strains belonging to several groups in the CFB complex play an important role in the normal development of green algae in the marine coastal environment.     

Protection by natural blackwater against disturbances in ion fluxes caused by low pH exposure in freshwater stingrays endemic to the Rio Negro

Stenohaline freshwater stingrays (Potamotrygon spp.) are endemic to the very dilute (Na(+), Cl(-), Ca2(+) 300 micromol L(-1) in reference water (low DOC) to about 100 micromol L(-1) in blackwater (high DOC). In reference water, both JNain and JClin were inhibited >90%, both JNaout and JClout more than doubled, and J(Amm) did not change at pH 4.0. In blackwater, the inhibition of influxes was attenuated, the increases in outflux did not occur, and J(Amm) increased by 60% at pH 4.0. Addition of 100 micromol L(-1) Ca(2+) to reference water prevented the increases in JNaout and JClout and allowed J(Amm) to increase at pH 4.0, which demonstrates that the gills are sensitive to Ca(2+). However, addition of Ca(2+) to blackwater had no effect on the responses to pH 4.0. Addition of commercial humic acid to reference water did not duplicate the effects of natural Rio Negro blackwater at the same DOC level; instead, it greatly exacerbated the increases in JNaout and JClout at low pH and prevented any protective influence of added Ca(2+). Thus, blackwater DOC appears to be very different from commercial humic acid. Biogeochemical modeling indicated that blackwater DOC prevents Ca(2+) binding, but not H(+) binding, to the gills and that the protective effects of blackwater cannot be attributed to its higher buffer capacity or its elevated Al or Fe levels. Natural DOC may act directly at the gills at low pH to exert a protective effect and, when doing so, may override any protective action of Ca(2+) that might otherwise occur.