Extreme low pH, not Al3+ , is a key abiotic stressor for the extremophyte Carex angustisquama (Cyperaceae) in highly acidic solfatara fields.

Volcanic acidification creates extreme soil conditions, where rhizotoxicity from extremely low pH (2-3) and high Al3+ strongly inhibit plant growth. C. angustisquama is a dominant extremophyte in highly acidic solfatara fields, where no other vascular plants can survive. Here we investigated the key abiotic stressor determining survival of this extremophyte. Soil analyses and topographic surveys were conducted to examine the effects of low pH and Al3+ , two major abiotic stressors in acidic soils, on the occurrence of C. angustisquama in solfatara fields. Hydroponic culture experiments were also performed to test its growth responses to these stressors. In field surveys, the spatial distribution of soil pH was consistent with vegetation zonation within a solfatara field. In contrast, soil exchangeable Al content was overall low due to strong eluviation. Statistical analysis also supported the significant role of soil pH in determining the distribution of C. angustisquama in a solfatara field. Furthermore, hydroponic culture experiments revealed a higher tolerance of C. angustisquama to low pH than a sister species, especially in the range pH 2-3, corresponding to the pH values of the actual habitats of C. angustisquama. Conversely, no significant interspecific difference was detected in Al3+ tolerance, indicating that both species had high Al3+ tolerance. This study suggests that low pH is a critical abiotic stressor leading to formation of the extremophyte in highly acidic solfatara fields. In contrast, C. angustisquama displayed high tolerance to Al3+ toxicity, probably acquired prior to speciation.

Comparative reproductive ecology of two sister Asarum species (Aristolochiaceae) in relation to the evolution of elongated floral appendage.

Genus Asarum (Aristolochiaceae) shows diverse floral morphology and is hypothesized to have diversified as a result of pollinator-mediated selection. Yet most aspects of their reproductive ecology, including pollinators, remain unclear. This study focuses on A. costatum and A. minamitanianum in Japan, a sister species pair having remarkable differences in calyx lobe length (10-20 mm and 70-180 mm, respectively). The objectives of this study are to elucidate multiple aspects of reproductive ecology of these two species and obtain evolutionary insights into floral organ elongation. We adopted combined approaches, including field observations, molecular analyses and cultivation experiments, such as pollinator observation for 3 years, fine-scale spatial genetic analysis of 769 individuals, paternity analysis based on 566 seeds over 4 years, and control pollination experiments. Both Asarum species had strong spatial genetic structures, indicating limited seed dispersal. Pollinator observation revealed that flies and ground-dwelling insects visited flowers of both species, but that the pollinator fauna differed between the species. The visitation rate of flies was extremely low but was more than twice as high in the species with an elongated floral appendage. Paternity analysis revealed A. minamitanianum was predominantly outcrossing, while A. costatum showed a wide range of selfing rates among fruits. These two Asarum species are likely adapted to fly pollination in the shady forest understorey, where available pollinator fauna is limited. In addition, although its function remains unclear, the elongated calyx lobe of A. minamitanianum could have evolved for effective pollen dispersal by attracting fly visitors.

Approximate Bayesian computation analysis of EST-associated microsatellites indicates that the broadleaved evergreen tree Castanopsis sieboldii survived the Last Glacial Maximum in multiple refugia in Japan.

Climatic changes have played major roles in plants' evolutionary history. Glacial oscillations have been particularly important, but some of their effects on plants' populations are poorly understood, including the numbers and locations of refugia in Asian warm temperate zones. In the present study, we investigated the demographic history of the broadleaved evergreen tree species Castanopsis sieboldii (Fagaceae) during the last glacial period in Japan. We used approximate Bayesian computation (ABC) for model comparison and parameter estimation for the demographic modeling using 27 EST-associated microsatellites. We also performed the species distribution modeling (SDM). The results strongly support a demographic scenario that the Ryukyu Islands and the western parts in the main islands (Kyushu and western Shikoku) were derived from separate refugia and the eastern parts in the main islands and the Japan Sea groups were diverged from the western parts prior to the coldest stage of the Last Glacial Maximum (LGM). Our data indicate that multiple refugia survived at least one in the Ryukyu Islands, and the other three regions of the western and eastern parts and around the Japan Sea of the main islands of Japan during the LGM. The SDM analysis also suggests the potential habitats under LGM climate conditions were mainly located along the Pacific Ocean side of the coastal region. Our ABC-based study helps efforts resolve the demographic history of a dominant species in warm temperate broadleaved forests during and after the last glacial period, which provides a basic model for future phylogeographical studies using this approach.

Importance of demographic history for phylogeographic inference on the arctic-alpine plant Phyllodoce caerulea in East Asia.

Arctic-alpine plants have enormous ranges in the Northern Hemisphere. Phylogeographic studies have provided insights into their glacial survival as well as their postglacial colonization history. However, our understanding of the population dynamics of disjunct alpine populations in temperate regions remains limited. During Pleistocene cold periods, alpine populations of arctic-alpine species in East Asia were either connected to an ice-free Beringia refugium or they persisted with prolonged isolation after their establishment. To estimate which of these scenarios is more likely, we elucidated the genetic structure of Phyllodoce caerulea (Ericaceae) in Beringia and northern Japan, East Asia. Sequence variation in multiple nuclear loci revealed that P. caerulea can be distinguished into northern and southern groups. A demographic analysis demonstrated that the north-south divergence did not predate the last glacial period and detected introgression from Phyllodoce aleutica, relative widely distributed in East Asia, exclusively into the southern group. Therefore, although there has been genetic divergence between northern Japan and Beringia in P. caerulea, the divergence is unlikely to have resulted from their prolonged geographic separation throughout several cycles of glacial and interglacial periods. Instead, our study suggests that the introgression contributed to the genetic divergence of P. caerulea and that the range of P. caerulea was plausibly connected between northern Japan and Beringia during the last glacial period. Overall, our study not only provides a biogeographic insight into alpine populations of arctic-alpine plants in East Asia but also emphasizes the importance of careful interpretation of genetic structure for inferring phylogeographic history.

Isolation and characterization of microsatellite loci in Ainsliaea faurieana (Asteraceae), an endemic plant species on Yakushima Island, Japan, and cross-species amplification in closely related taxa.

Fourteen microsatellite loci were isolated and characterized for Ainsliaea faurieana, an endemic perennial plant species on Yakushima Island. In an analysis of 37 individuals from Yakushima Island, the observed number of alleles ranged from two to six. The expected and observed heterozygosities were 0.109-0.811 and 0.000-0.27, respectively. The inbreeding coefficient was 0.489-1.000. The interspecific applicability of these microsatellites was evaluated by analysing three closely related taxa from the Ryukyu Islands. All primer pairs for the 14 loci tested successfully amplified in all taxa.

Leukotriene C(4) enhances the contraction of porcine tracheal smooth muscle through the activation of Y-27632, a rho kinase inhibitor, sensitive pathway.

1. An unsaturated fatty acid, leukotriene C(4) (LTC(4)), has a potent contractile effect on human airway smooth muscle, and has been implicated in the pathogenesis of human asthma. Using front-surface fluorometry with fura-PE3, the effect of LTC(4) on the intracellular Ca(2+) concentration ([Ca(2+)](i)) and tension were investigated in porcine tracheal smooth muscle strips. 2. The application of LTC(4) induced little or no contraction despite a small and transient increase in [Ca(2+)](i). In the presence of LTC(4), however, the contractions evoked by high K(+) depolarization or a low concentration of carbachol (CCh) were markedly enhanced without inducing any changes in the [Ca(2+)](i) levels, thus indicating that LTC(4) increases the Ca(2+) responsiveness of the contractile apparatus. This LTC(4)-induced increase in Ca(2+) responsiveness could partly be reproduced in the permeabilized preparation of tracheal smooth muscle strips. 3. The LTC(4)-induced enhancement of contraction was accompanied by an increase in myosin light chain (MLC) phosphorylation and was blocked by a rho kinase inhibitor (Y-27632), but not by either a PKC inhibitor (calphostin C) or a tyrosine kinase inhibitor (genistein). 4. These results indicated that, in porcine tracheal smooth muscle, LTC(4) enhances the contraction by increasing the Ca(2+) responsiveness of the contractile apparatus in a MLC phosphorylation dependent manner, possibly through the activation of the rho-rho kinase pathway.

[Regulation of vascular tone by the modulation of Ca2+ sensitivity].

The contractile tone of the vascular smooth muscle plays an important role on the regulation of the blood pressure as well as the local perfusion of the important organs such as the heart and brain. The importance of the Ca(2+) sensitivity in the regulation of the vascular tone has been established by the development of the simultaneous measurements of intracellular Ca(2+) concentration ( [Ca(2+)](i) ) and tension as well as that of the receptor coupled permeabilized preparation in the late 1980s. Recently, the mechanisms underlying the regulation of Ca(2+) sensitivity have been revealed. The increase in the Ca(2+) sensitivity involves the myosin phosphatase (MLCP) inhibition mediated by rhoA-rho kinase system and PKC-CPI system. The decrease in the Ca(2+) sensitivity involves the PKA-mediated inhibition of myosin light chain kinase, the PKG-mediated activation of MLCP, and PKA- or PKG-mediated inactivation of rhoA. In this article, the regulation of the Ca(2+) sensitivity of the contractile apparatus of the vascular smooth muscle will be briefly reviewed.

Intersectional gene flow between insular endemics of Ilex (Aquifoliaceae) on the Bonin Islands and the Ryukyu Islands.

Hybridization and introgression play important roles in plant evolution, and their occurrence on the oceanic islands provides good examples of plant speciation and diversification. Restriction fragment length polymorphisms (RFLPs) and trnL (UAA) 3'exon-trnF (GAA) intergenic spacer (IGS) sequences of chloroplast DNA (cpDNA), and the sequences of internal transcribed spacer (ITS) of nuclear ribosomal DNA were examined to investigate the occurrence of gene transfer in Ilex species on the Bonin Islands and the Ryukyu Islands in Japan. A gene phylogeny for the plastid genome is in agreement with the morphologically based taxonomy, whereas the nuclear genome phylogeny clusters putatively unrelated endemics both on the Bonin and the Ryukyu Islands. Intersectional hybridization and nuclear gene flow were independently observed in insular endemics of Ilex on both sets of islands without evidence of plastid introgression. Gene flow observed in these island systems can be explained by ecological features of insular endemics, i.e., limits of distribution range or sympatric distribution in a small land area.

Phylogenetic relationships within Araucariaceae based on rbcL gene sequences.

Phylogenetic relationships were determined in the Araucariaceae, which are now found mainly in the Southern Hemisphere. This conifer family was well diversified and widely distributed in both hemispheres during the Mesozoic era. The sequence of 1322 bases of the rbcL gene of cpDNA was determined from 29 species of Araucariaceae, representing almost all the species of the family. Phylogenetic trees determined by the parsimony method indicate that Araucariaceae are well defined by rbcL sequences and also that the monophyly of Agathis or Araucaria is well supported by high bootstrap values. The topology of these trees revealed that Wollemia had derived prior to Agathis and Araucaria. The rbcL phylogeny agrees well with the present recognition of four sections within Araucaria: Araucaria, Bunya, Eutacta, and Intermedia. Morphological characteristics of the number of cotyledons, position of male cone, and cuticular micromorphologies were evaluated as being phylogenetically informative. Section Bunya was found to be derived rather than to be the oldest taxon. Infrageneric relationships of Agathis could not be well elucidated because there are few informative site changes in the rbcL gene, suggesting the more recent differentiation of the species as their fossil records indicate. The New Caledonian Araucaria and Agathis species each formed a monophyletic group with very low differentiation in rbcL sequences among them, indicating rapid adaptive radiation to new edaphic conditions, i.e., ultramafic soils, in the post-Eocene era.