The hagfish genome and the evolution of vertebrates.

As the only surviving lineages of jawless fishes, hagfishes and lampreys provide a crucial window into early vertebrate evolution1-3. Here we investigate the complex history, timing and functional role of genome-wide duplications4-7 and programmed DNA elimination8,9 in vertebrates in the light of a chromosome-scale genome sequence for the brown hagfish Eptatretus atami. Combining evidence from syntenic and phylogenetic analyses, we establish a comprehensive picture of vertebrate genome evolution, including an auto-tetraploidization (1RV) that predates the early Cambrian cyclostome-gnathostome split, followed by a mid-late Cambrian allo-tetraploidization (2RJV) in gnathostomes and a prolonged Cambrian-Ordovician hexaploidization (2RCY) in cyclostomes. Subsequently, hagfishes underwent extensive genomic changes, with chromosomal fusions accompanied by the loss of genes that are essential for organ systems (for example, genes involved in the development of eyes and in the proliferation of osteoclasts); these changes account, in part, for the simplification of the hagfish body plan1,2. Finally, we characterize programmed DNA elimination in hagfish, identifying protein-coding genes and repetitive elements that are deleted from somatic cell lineages during early development. The elimination of these germline-specific genes provides a mechanism for resolving genetic conflict between soma and germline by repressing germline and pluripotency functions, paralleling findings in lampreys10,11. Reconstruction of the early genomic history of vertebrates provides a framework for further investigations of the evolution of cyclostomes and jawed vertebrates.

Ontogenetic Expression of Aquaporins in the Kidney and Urinary Bladder of the Japanese Tree Frog, Dryophytes japonicus.

For adult anuran amphibians, the kidney and urinary bladder play important osmoregulatory roles through water reabsorption. In the present study, we have examined ontogenetic expression of aquaporins, i.e., AQP2, AQPamU (AQP6ub, AQPa2U), and AQP3, in these organs using the Japanese tree frog, Dryophytes japonicus. Immunohistochemistry using the metamorphosing larvae at stages 40-43 localized AQP2 protein to the collecting ducts in the dorsal zone of the mesonephric kidney. At prometamorphic stages 40 and 41, labelling of AQP2 protein was observed in the apical/ subapical regions of the collecting duct cells. At climax stages 42 and 43, labels for AQP2 and AQP3 became observed in the apical/subapical regions and basolateral membrane of the collecting duct cells, respectively, as seen in the adults. As for the urinary bladder, immuno-positive labels for AQPamU were localized to the apical/subapical regions of granular cells in the mucosal epithelium at stages 40-43. On the other hand, AQP3 immunoreactivity was hardly observed in the urinary bladder at stage 40, and weakly appeared in many granular cells at stage 41. Thereafter, labels for AQP3 became evident along the basolateral membrane of granular cells at stages 42 and 43, together with AQPamU in the apical/subapical regions. These results suggest that the kidney and urinary bladder might be capable of water reabsorption, via AQP2, AQPamU, and AQP3, at stage 42, contributing to the acclimation of the tree frogs to terrestrial environments.

A novel type of prolactin expressed in the bullfrog pituitary specifically during the larval period.

Prolactin (PRL) is one of the major hormones that control amphibian metamorphosis. Recently, a PRL (PRL1B) gene that is different from the known PRL (PRL1A) gene has been found in the genomes of several amphibian species. In order to ascertain whether the PRL1B gene is expressed in the bullfrog (Rana catesbeiana) pituitary, cloning of cDNA encoding PRL1B in the pituitary of the premetamorphic bullfrog tadpole was attempted. The bullfrog PRL1B amino acid sequence predicted from the obtained cDNA showed 62% identity with those of Xenopus PRL1Bs that have been presumed from the genome sequences, whereas the sequence identity between bullfrog PRL1A and PRL1B was 48%. A molecular phylogenetic tree showed that bullfrog PRL1B is most appropriately grouped with amphibian PRL1Bs. Quantitative PCR analysis revealed that the mRNA expression levels of bullfrog PRL1B in the pituitary were high during pre- and prometamorphosis, sharply declined at metamorphic climax and became undetectable after metamorphosis. In contrast, PRL1A mRNA levels were relatively low during pre- and prometamorphosis, rose at climax and remained high after metamorphosis. Immunohistochemical study using antibodies against partial peptides of PRL1A and PRL1B revealed that most of the PRL1A- and PRL1B-immunoreactive cells in the larval pituitary were distributed separately, but that some of the cells immunoreactive with both antibodies were also present. Western blot analysis with the larval pituitary extract indicated that PRL1B-immunoreactive band appeared at the position of molecular weight ca. 22.1 kDa and PRL1A-immunoreactive band at the position of ca. 22.8 kDa. The results obtained in this experiment suggest the possibility that PRL1B plays as-yet-unknown role(s) during the pre-climactic period of metamorphosis. This is the first report on the existence of PRL1B as a protein in the amphibian larval pituitary.

Molecular Cloning of cDNA Encoding an Aquaglyceroporin, AQP-h9, in the Japanese Tree Frog, Hyla japonica: Possible Roles of AQP-h9 in Freeze Tolerance.

In order to study the freeze-tolerance mechanism in the Japanese tree frog, Hyla japonica, wecloned a eDNA encoding aquaporin (AQP) 9 from its liver. The predicted amino acid sequence ofH. japonica AQP9 (AQP-h9) contained six putative transmembrane domains and two conservedAsn-Pro-Aia motifs, which are characteristic of AQPs. A swelling assay using Xenopus laevisoocytes injected with AQP-h9 cRNA showed that AQP-h9 facilitated water and glycerol permeation,confirming its property as an aquaglyceroporin. Subsequently, glycerol concentrations in serumand tissue extracts were compared among tree frogs that were hibernating, frozen, or thawed afterfreezing. Serum glycerol concentration of thawed frogs was significantly higher than that of hibernatingfrogs. Glycerol content in the liver did not change in the freezing experiment, whereas thatin the skeletal muscle was elevated in thawed frogs as compared with hibernating or frozen frogs. Histological examination of the liver showed that erythrocytes aggregated in the sinusoids during hibernation and freezing, and immunoreactive AQP-h9 protein was detected over the erythrocytes. The AQP-h9 labeling was more intense in frozen frogs than in hibernating frogs, but nearly undetectable in thawed frogs. For the skeletal muscle, weak labels for AQP-h9 were observed in the cytoplasm of myocytes of hibernating frogs. AQP-h9 labeling was markedly enhanced by freezing and was decreased by thawing. These results indicate that glycerol may act as a c;:ryoprotectant in H. japonica and that during hibernation, particularly during freezing, AQP-h9 may be involved in glycerol uptake in erythrocytes in the liver and in intracellular glycerol transport in the skeletal muscle cells.

Molecular and cellular characterization of urinary bladder-type aquaporin in Xenopus laevis.

In contrast to many anuran amphibians, water is not reabsorbed from the urinary bladder in aquatic Xenopus, thereby helping to prevent excessive water influx. However, little is known about the molecular mechanisms for this process. In the present study, we have identified urinary bladder-type aquaporin, AQP-x2, in Xenopus laevis by cDNA cloning. The predicted amino acid sequence contained six putative transmembrane domains and the two conserved Asn-Pro-Ala motifs, characteristic of AQPs. The sequence also contained a putative N-glycosylation site and phosphorylation motifs for protein kinase A and protein kinase C. The oocyte swelling assay showed that AQP-x2 facilitated water permeability. Reverse transcription-PCR analysis indicated that AQP-x2 mRNA was expressed in the urinary bladder and lung, and faintly in the kidney. Immunomicroscopical study further localized AQP-x2 protein to the cytoplasm of granular cells in the luminal epithelium of the urinary bladder whilst AQP3 was observed along the basolateral side of these cells. In vitro stimulation of the urinary bladder with 10(-8)M vasotocin (AVT), 10(-8)M hydrin 1, or 10(-8)M hydrin 2 had no clear effect on the subcellular distribution of AQP-x2. When the AVT concentration was increased to 10(-6)M, however, AQP-x2 was partially transferred to the apical plasma membrane. The treatment with hydrin 1 or hydrin 2 at the same concentration failed to induce the translocation to the apical membrane. On the other hand, AQP3 remained along the basolateral side even after the treatment with vasotocin or hydrins. The results suggest that the poor responsiveness of AQP-x2 to neurohypophyseal peptides may be a main cause for the little water permeability of the urinary bladder of X. laevis.

Throughfall under a teak plantation in Thailand: a multifactorial analysis on the effects of canopy phenology and meteorological conditions.

Valuable teak (Tectona grandis Linn. f.) plantations cover vast areas throughout Southeast Asia. This study sought to increase our understanding of throughfall inputs under teak by analyzing the abiotic and biotic factors governing throughfall amounts and ratios in relation to three canopy phenophases (leafless, leafing, and leafed). There was no rain during the brief leaf senescence phenophase in our study. Leveraging detailed field observations, we employed boosted regression tree (BRT) analysis to identify the primary controls on throughfall amount and ratio during each canopy phenophase. Whereas throughfall amounts were always dominated by rainfall magnitude (as expected), throughfall ratios were governed by a suite of predictor variables during each phenophase. The BRT analysis demonstrated that throughfall ratio in the leafless phase was most influenced (in descending order of importance) by air temperature, rainfall amount, maximum wind speed, and rainfall intensity. Throughfall ratio in the leafed phenophase was dominated by rainfall amount. The leafing phenophase was an intermediate case where rainfall amount, air temperature, and vapor pressure deficit were most important. Our results highlight the fact that throughfall ratios are differentially influenced by a suite of meteorological variables during each canopy phenophase. Abiotic variables, such as rainfall amount and air temperature, trumped leaf area index and stand density in their effect on throughfall ratio. The leafing phenophase, while transitional in nature and short in duration, has a detectable and unique impact on water inputs to teak plantations. Further work is needed to better understand the biogeochemistry of leaf emergence in teak plantations.

Injectable cell scaffold restores impaired cell-based therapeutic angiogenesis in diabetic mice with hindlimb ischemia.

The clinical success of cell-based therapeutic angiogenesis has been limited in diabetic patients with critical limb ischemia. We previously reported that an injectable cell scaffold (ICS), which is a nano-scaled hydroxyapatite (HAp)-coated polymer microsphere, enhances therapeutic angiogenesis. Subsequently, we developed a modified ICS for clinical use, measuring 50 µm in diameter using poly(l-lactide-co-ε-caprolactone) as a biodegradable polymer, which achieved appropriately accelerated absorption in vivo. The aim of the present study was to evaluate the effectiveness of this practical ICS in diabetic hindlimb ischemia. Bone-marrow mononuclear cells (BMNCs) were intramuscularly injected, without or with a practical ICS, into the ischemic hindlimbs of mice (BMNCs or ICS+BMNCs group, respectively). Kaplan-Meier analysis demonstrated that the beneficial effects of BMNC transplantation for limb salvage after ischemic surgery were almost entirely abrogated in streptozotocin-induced diabetic mice. In contrast, injection of ICS+BMNCs revealed significant limb salvage in diabetic mice to a similar extent as in non-diabetic mice. The number of apoptotic transplanted BMNCs was 1.8-fold higher in diabetic mice 10 days after transplantation compared to non-diabetic mice, while that in the ICS+BMNCs group was markedly lower (8.3% of that in the BMNCs group) even in diabetic mice. The proangiogenic factors VEGF and FGF2, also known as antiapoptotic factors, mostly co-localized with transplanted GFP-positive BMNCs that were closely aggregated around the ICS in ischemic tissue. In conclusion, the practical ICS significantly augmented cell-based therapeutic angiogenesis even in diabetic animals, through local accumulation of proangiogenic factors and antiapoptotic effects in transplanted cells.

Gene expression and localization of two types of AQP5 in Xenopus tropicalis under hydration and dehydration.

Two types of aquaporin 5 (AQP5) genes (aqp-xt5a and aqp-xt5b) were identified in the genome of Xenopus tropicalis by synteny comparison and molecular phylogenetic analysis. When the frogs were in water, AQP-xt5a mRNA was expressed in the skin and urinary bladder. The expression of AQP-xt5a mRNA was significantly increased in dehydrated frogs. AQP-xt5b mRNA was also detected in the skin and increased in response to dehydration. Additionally, AQP-xt5b mRNA began to be slightly expressed in the lung and stomach after dehydration. For the pelvic skin of hydrated frogs, immunofluorescence staining localized AQP-xt5a and AQP-xt5b to the cytoplasm of secretory cells of the granular glands and the apical plasma membrane of secretory cells of the small granular glands, respectively. After dehydration, the locations of both AQPs in their respective glands did not change, but AQP-xt5a was visualized in the cytoplasm of secretory cells of the small granular glands. For the urinary bladder, AQP-xt5a was observed in the apical plasma membrane and cytoplasm of a number of granular cells under normal hydration. After dehydration, AQP-xt5a was found in the apical membrane and cytoplasm of most granular cells. Injection of vasotocin into hydrated frogs did not induce these changes in the localization of AQP-xt5a in the small granular glands and urinary bladder, however. The results suggest that AQP-xt5a might be involved in water reabsorption from the urinary bladder during dehydration, whereas AQP-xt5b might play a role in water secretion from the small granular gland.

Nitrate isotopic composition reveals nitrogen deposition and transformation dynamics along the canopy-soil continuum of a suburban forest in Japan.

RATIONALE: Heavy nitrogen (N) deposition often causes high nitrate (NO3(-)) accumulation in soils in temperate forested ecosystems. To clarify the sources and production pathways of this NO3(-), we investigated NO3(-) isotope signatures in deposition processes along the canopy-soil continuum of a suburban forest in Japan. METHODS: The stable isotopes of N and oxygen (O) were used to trace the source and transformation dynamics of nitrate (NO3(-)) in two forest stands: a plantation of Cryptomeria japonica (coniferous tree; CJ) and a natural secondary forest of Quercus acutissima (broadleaf, deciduous tree; QA). The NO3(-) and ammonium (NH4(+)) concentrations were measured, as well as the δ(15)N and δ(18)O values of NO3(-), in rainfall, throughfall, stem flow, litter layer water, and soil water (10, 30, and 70 cm depths). RESULTS: Seasonal variations were observed in the δ(15)N values of throughfall and stem flow NO3(-) at both sites, and in the δ(18)O values of throughfall and stem flow NO3(-) at the QA site. The range in the δ(18)O values of rainfall and throughfall NO3(-) was large (65-70‰) but decreased dramatically to 2-5‰ in soil water at both sites. At the QA site, the δ(18)O values of stem flow NO3(-) decreased to 40‰ during several rain events, especially in the growing season. CONCLUSIONS: NO3(-) from atmospheric deposition was replaced by microbially generated NO3(-) mainly in the organic horizon and surface portion of the mineral soil under excess N deposition in this suburban forest. Microbial activity, including both immobilization and nitrification in organic-rich horizons near the surface, contributed to incorporating atmospheric NO3(-) quickly into the internal microbial N cycle. We also found evidence of microbial nitrification in the canopy of the QA stand during the growing season.

Molecular cloning and functional characterization of two forms of Pax8 in the rainbow trout, Oncorhynchus mykiss.

We have identified two distinct Pax8 (a and b) mRNAs from the thyroid gland of the rainbow trout (Oncorhynchus mykiss), which seemed to be generated by alternative splicing. Both Pax8a and Pax8b proteins were predicted to possess the paired domain, octapeptide, and partial homeodomain, while Pax8b lacked the carboxy-terminal portion due to an insertion in the coding region of the mRNA. RT-PCR analysis showed each of Pax8a and Pax8b mRNAs to be abundantly expressed in the thyroid and kidney. In situ hybridization histochemistry further detected the expression of Pax8 mRNA in the epithelial cells of the thyroid follicles of the adult trout and in the thyroid primordial cells of the embryo. The functional properties of Pax8a and Pax8b were investigated by dual luciferase assay. The transcriptional regulation by the rat thyroid peroxidase (TPO) promoter was found to be increased by Pax8a, but not by Pax8b. Pax8a further showed synergistic transcriptional activity with rat Nkx2-1 for the human TPO upstream region including the enhancer and promoter. On the other hand, Pax8b decreased the synergistic activity of Pax8a and Nkx2-1. Electrophoretic mobility shift assay additionally indicated that not only Pax8a but also Pax8b can bind to the TPO promoter and enhancer, implying that the inhibitory effect of Pax8b might result from the lack of the functional carboxy-terminal portion. Collectively, the results suggest that for the trout thyroid gland, Pax8a may directly increase TPO gene expression in cooperation with Nkx2-1 while Pax8b may work as a non-activating competitor for the TPO transcription.

Gene expression, function, and diversity of Nkx2-4 in the rainbow trout, Oncorhynchus mykiss.

Nkx2 homeodomain transcription factors are involved in various developmental processes and cell specification: e.g. in mammals, NKX2-1 is essential for thyroid-specific gene expression and thyroid morphogenesis. Among Nkx2 proteins, information is still very limited for Nkx2-4. In the present study, we have identified three distinct cDNAs encoding Nkx2-4 isoforms (Nkx2-4a, -b, and -c) from the rainbow trout thyroid tissue, and characterized their transcriptional properties. The trout Nkx2-4 proteins were all predicted to conserve three characteristic domains: the tinman-like amino terminal decapeptide, the NK2 homeodomain, and the NK2-specific domain, and also share 75-89% amino acid similarity. It was shown by dual luciferase assay that Nkx2-4a and Nkx2-4b, but not Nkx2-4c, significantly activated transcription from a cotransfected rat thyroglobulin (TG) promoter. An electrophoretic mobility shift assay indicated that all the Nkx2-4 isoforms could bind to the TG promoter, implying that the faint transcriptional activity of Nkx2-4c might result from some critical amino acid substitution(s) outside the homeodomain. RT-PCR analysis revealed similar tissue distribution patterns for Nkx2-4a and Nkx2-4b mRNAs. Both mRNAs were expressed abundantly in the thyroid, and weakly in the testis. On the other hand, Nkx2-4c mRNA was detected in the ovary as well as in the thyroid. The expression sites of Nkx2-4c mRNA were localized, by in situ hybridization histochemistry, to the ovarian granulosa cells and to the thyroid follicular cells. The results suggest that in the rainbow trout, Nkx2-4a and Nkx2-4b might play a major role in TG gene transcription whereas Nkx2-4c might have some functions in the ovary as well as the thyroid.

Inter-annual variation in the response of leaf-out onset to soil moisture increase in a teak plantation in northern Thailand.

To understand the impact of inter-annual climate change on vegetation-atmosphere mass and energy exchanges, it has become necessary to explore changes in leaf-out onset in response to climatic fluctuations. We examined the response of leaf-out and transpiration onset dates to soil moisture in a teak plantation in northern Thailand based on a 12-year leaf area index and sap flow measurements. The date of leaf-out and transpiration onset varied between years by up to 40 days, and depended on the initial date when the relative extractable water in a soil layer of 0-0.6 m (Θ) was greater than 0.2 being consistent with our previous results. Our new finding is that the delay in leaf-out and transpiration onset relative to the initial date when Θ > 0.2 increases linearly as the initial date on which Θ > 0.2 becomes earlier. The delay spans about 20 days in years when Θ > 0.2 occurs in March (the late dry season)-much earlier than usual because of heavy pre-monsoon rainfalls-while there is little delay in years when Θ > 0.2 occurs in May. This delay indicates the influence of additional factors on leaf-out onset, which controls the delay in the response of leaf-out to soil moisture increase. The results increased our knowledge about the pattern and extent of the changes in leaf phenology that occur in response to the inter-annual climate variation in tropical regions, where, in particular, such research is needed.

Gross anatomy of the Pacific hagfish, Eptatretus burgeri, with special reference to the coelomic viscera.

Hagfish (Myxinoidea) are a deep-sea taxon of cyclostomes, the extant jawless vertebrates. Many researchers have examined the anatomy and embryology of hagfish to shed light on the early evolution of vertebrates; however, the diversity within hagfish is often overlooked. Hagfish have three lineages, Myxininae, Eptatretinae, and Rubicundinae. Usually, textbook illustrations of hagfish anatomy reflect the morphology of the Myxininae lineage, especially Myxine glutinosa, with its single pair of external branchial pores. Here, we instead report the gross anatomy of an Eptatretinae, Eptatretus burgeri, which has six pairs of branchial pores, especially focusing on the coelomic organs. Dissections were performed on fixed and unfixed specimens to provide a guide for those doing organ- or tissue-specific molecular experiments. Our dissections revealed that the ventral aorta is Y-branched in E. burgeri, which differs from the unbranched morphology of Myxine. Otherwise, there were no differences in the morphology of the lingual apparatus or heart in the pharyngeal domain. The thyroid follicles were scattered around the ventral aorta, as has been reported for adult lampreys. The hepatobiliary system more closely resembled those of jawed vertebrates than those of adult lampreys, with the liver having two lobes and a bile duct connecting the gallbladder to each lobe. Overall, the visceral morphology of E. burgeri does not differ significantly from that of the known Myxine at the level of gross anatomy, although the branchial morphology is phylogenetically ancestral compared to Myxine.

Angiogenesis in the intermediate lobe of the pituitary gland alters its structure and function.

The pars distalis (PD) and the pars intermedia (PI) have the same embryonic origin, but their morphological and functional characteristics diverge during development. The PD is highly vascularized, whereas the highly innervated PI is essentially non-vascularized. Based on our previous finding that vascular endothelial growth factor-A (VEGF-A) is involved in vascularization of the rat PD, attempt was made to generate transgenic Xenopus expressing VEGF-A specifically in the melanotrope cells of the PI as a model system for studying the significance of vascularization or avascularization for the functional differentiation of the pituitary. The PI of the transgenic frogs, examined after metamorphosis, were distinctly vascularized but poorly innervated. The experimentally induced vascularization in the PI resulted in a marked increase in tissue volume and a decrease in the expression of both alpha-melanophore-stimulating hormone (α-MSH) and prohormone convertase 2, a cleavage enzyme essential for generating α-MSH. The transgenic animals had low plasma α-MSH concentrations and displayed incomplete adaptation to a black background. To our knowledge, this is the first report indicating that experimentally induced angiogenesis in the PI may bring about functional as well as structural alterations in this tissue.

Carbon allocation in a Bornean tropical rainforest without dry seasons.

To clarify characteristics of carbon (C) allocation in a Bornean tropical rainforest without dry seasons, gross primary production (GPP) and C allocation, i.e., above-ground net primary production (ANPP), aboveground plant respiration (APR), and total below-ground carbon flux (TBCF) for the forest were examined and compared with those from Amazonian tropical rainforests with dry seasons. GPP (30.61 MgC ha(-1) year(-1), eddy covariance measurements; 34.40 MgC ha(-1) year(-1), biometric measurements) was comparable to those for Amazonian rainforests. ANPP (6.76 MgC ha(-1) year(-1)) was comparable to, and APR (8.01 MgC ha(-1) year(-1)) was slightly lower than, their respective values for Amazonian rainforests, even though aboveground biomass was greater at our site. TBCF (19.63 MgC ha(-1) year(-1)) was higher than those for Amazonian forests. The comparable ANPP and higher TBCF were unexpected, since higher water availability would suggest less fine root competition for water, giving higher ANPP and lower TBCF to GPP. Low nutrient availability may explain the comparable ANPP and higher TBCF. These data show that there are variations in C allocation patterns among mature tropical rainforests, and the variations cannot be explained solely by differences in soil water availability.

On Support for Dyslexic Students in Senior-High School and Higher Education.

It is very difficult to provide dyslexic students in senior-high school or higher education with all necessary textbooks in ordinary accessible format such as multimedia DAISY. Here, a new approach to provide them with a new type of accessible textbooks named "Fixed-Layout DAISY" is shown. In it, the whole page is treated as a multi-layer picture, the front layer of which has the same form as the original PDF. A DAISY (EPUB3) player can read out any texts together with highlighting them. It does not have the reflow function. The page layout is always kept as same as the original. It does not have information either in which order texts on the page should be read out, and readers need to click a text block on a page where they want to read. Dyslexic people can see and click a place where they want to read, and obviously, Fixed-Layout DAISY should work for them. Fixed-Layout DAISY can be produced almost automatically from an original "e-born PDF" by making use of our OCR system, and it should be very helpful for the dyslexic students to get accessible version of their textbooks.

Multilingual Support for Accessibility in PowerPoint STEM Contents.

Many online educational materials for sighted students such as presentation slides, PDF materials, MP4 videos, etc. are produced with Microsoft PowerPoint (PPT). In terms of non-technical contents, accessible PPT contents can be produced in a certain level; however, as far as STEM contents are concerned, there is still a serious difficulty. Our new add-on for PPT allows users to make efficiently/easily PPT STEM contents accessible. By making use of it, alt text/aloud reading by a TTS voice can be added to any technical part such as mathematical expressions included in PPT slides. An accessible MP4 video for STEM education also can be produced efficiently. By making use of multilingual support in Infty software, this add-on has been recently improved so that users can use it for PPT contents in various local languages other than Japanese or English.

The hagfish genome and the evolution of vertebrates.

As the only surviving lineages of jawless fishes, hagfishes and lampreys provide a critical window into early vertebrate evolution. Here, we investigate the complex history, timing, and functional role of genome-wide duplications in vertebrates in the light of a chromosome-scale genome of the brown hagfish Eptatretus atami. Using robust chromosome-scale (paralogon-based) phylogenetic methods, we confirm the monophyly of cyclostomes, document an auto-tetraploidization (1RV) that predated the origin of crown group vertebrates ~517 Mya, and establish the timing of subsequent independent duplications in the gnathostome and cyclostome lineages. Some 1RV gene duplications can be linked to key vertebrate innovations, suggesting that this early genomewide event contributed to the emergence of pan-vertebrate features such as neural crest. The hagfish karyotype is derived by numerous fusions relative to the ancestral cyclostome arrangement preserved by lampreys. These genomic changes were accompanied by the loss of genes essential for organ systems (eyes, osteoclast) that are absent in hagfish, accounting in part for the simplification of the hagfish body plan; other gene family expansions account for hagfishes' capacity to produce slime. Finally, we characterise programmed DNA elimination in somatic cells of hagfish, identifying protein-coding and repetitive elements that are deleted during development. As in lampreys, the elimination of these genes provides a mechanism for resolving genetic conflict between soma and germline by repressing germline/pluripotency functions. Reconstruction of the early genomic history of vertebrates provides a framework for further exploration of vertebrate novelties.

Possible molecular mechanism for acute encephalopathy by angel-wing mushroom ingestion - Involvement of three constituents in onset.

In Japan in 2004, 59 people who had consumed angel-wing mushroom, Pleurocybella porrigens, experienced acute encephalopathy, and of these 17 died. We purified a lethal protein to mice, pleurocybelline (PC), from P. porrigens. Although PC caused no damage to the brain, PC formed a complex with a lectin (PPL) and showed exo-protease activity, degrading substrates from both N- and C-termini. In addition, the presence of an unstable toxic compound, pleurocybellaziridine (PA), in the mushroom was demonstrated. We hypothesized that the complex and PA are involved in disease development and verified that apoptotic cells in the hippocampus were significantly increased by injection of the mixture of PC, PPL, and PA, indicating that these substances might be involved in acute encephalopathy.

Mechanical stability of orthodontic miniscrew depends on a thread shape.

Background/purpose: Primary stability of orthodontic miniscrew system is of great importance in maintaining stable anchorage during a treatment period. Thus, this study aimed to examine whether the thread shape of orthodontic miniscrew had an effect on its mechanical stability in bone. Materials and methods: Three different types of miniscrews (type A and B with a regular thread shape; type C with a novel thread shape) were placed in artificial bone block with different artificial cortical bone thickness of 1.5, 2.0 and 3.0 mm. Values of maximum insertion torque (MIT), removal torque (RT), torque ratio (TR), screw mobility, static stiffness (K), dynamic stiffness (K∗) and energy dissipation (tan Î´) ability were assessed for each miniscrew system. Results: The MIT, RT, TR and K of type C miniscrew were significantly greater than those of type A and B miniscrews when the miniscrews were placed in the thinner artificial bone. Furthermore, the TR value of type C miniscrew was more than 1, indicating the MRT value was larger than the MIT value in the novel miniscrew. The values of K∗ and tan Î´ were almost similar among the three types of miniscrews. Conclusion: The miniscrew with a novel thread shape showed a higher initial stability compared to those with a regular thread shape. Thus, in order to obtain a sufficient initial stability, it is important to select the type of screw thread that is appropriate for the thickness of the cortical bone.