ERF is a unique transcription factor in higher plants with AP2-like DNA-binding domains. An ERF-like gene was isolated from Galegae orientalis, termed GoDREB. Sequence alignment showed that it shares high identity with other ERF family members in AP2 domain. Transient expression analysis indicated that GoDREB protein is localized in the nucleus. Quantitative RT-PCR analysis results showed that GoDREB is induced by a variety of abiotic stress, such as cold, dehydration, and high-salinity. Exogenous hormones, such as methyl jasmonate acid and salicylic acid, also up-regulate the expression of GoDREB. However, ABA did not induce the mRNA accumulation of GoDREB. These results implied that the GoDREB might play a role in these two hormones-dependent but not be involved in the ABA-dependent stress signaling pathway. Overexpression of GoDREB in transgenic tobacco plants resulted in higher tolerance to high salinity, osmotic and low-temperature stresses. These results suggest that GoDREB may play an essential role as an ERF transcription factor in regulation of stress-responsive signaling in G. orientalis.

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Mlo is a plant-specific gene family, which is known to show stress responses in various plants. To reveal the genetic characteristics of the Mlo family in wheat, we isolated wheat Mlo members from a database and studied their expression in young shoots and roots under salt and osmotic stress conditions. In an in silico investigation, we identified seven Mlo members in wheat and named them TaMlo-1~TaMlo-7. None of the wheat Mlo showed significant induction or reduction of their expression under salt or osmotic stress, but organ-specific expression was observed in several TaMlo members. TaMlo-1, TaMlo-2, and TaMlo-5 were constitutively expressed in both shoots and roots, but TaMlo-3 and TaMlo-4 showed root-specific expression, and TaMlo-7 showed dominant expression in shoots. TaMlo-6 was weakly expressed in both shoots and roots. Phylogenetic analysis classified the plant Mlo members into six classes; four of them were comprised of angiosperm Mlo members, and the remaining two consisted of fern and moss Mlo members. The seven wheat Mlo members were classified into four angiosperm Mlo classes, similar to those of Arabidopsis and rice, indicating that the formation of each of the Mlo classes preceded the divergence of dicots and monocots. The differentiation of the expressional patterns among the seven TaMlo members was not related to their phylogenetic classification. This result suggested that the organ specific expression of individual Mlo members occurred relatively recently in their evolution.

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The interaction between figs (Ficus, Moraceae) and fig-pollinating wasps (Chalcidoidea, Agaonidae) is one of the most specific mutualisms, and thus is a model system for studying coevolution and cospeciation. In this study we focused on figs and their associated fig-wasps found in the Ryukyu and Bonin (Ogasawara) Islands, Japan, because it has been suggested that breakdown in the specificity may occur in islands or at edge of a species’ distribution. We collected 136 samples of 15 native fig species and 95 samples of 13 associated fig-wasps from all major islands in the Ryukyu Islands, including two fig species and one fig-wasp species endemic to the Bonin Islands. We performed molecular phylogenetic analyses using plastid DNA and nuclear ITS sequences for the figs and nuclear 28S rRNA and mitochondrial COI genes for the fig-wasps to investigate the interspecific phylogenies and intraspecific variation within the mutualism. Our phylogenetic analyses using multiple samples per species show the single clade of each fig (except the Bonin endemic species) and fig-pollinating wasp species. Fig species belonging to the same subgenera formed well-supported clades in both plastid and ITS trees, except for the subgenus Urostigma. Likewise, fig wasps emerging from host fig species belonging to the same subgenera formed mostly well supported clades in both 28S and COI trees. Host specificity between the figs and fig-wasps functions strictly in these islands. There was very little sequence variation within species, and that no major geographic structure was found. The two Bonin endemic species (F. boninsimae and F. nishimurae) or their common ancestor and the associated fig-wasps (Blastophaga sp.) are apparently derived from F. erecta and its associated fig-wasps (B. nipponica), respectively, and probably migrated from the Ryukyu Islands.

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The Pax gene family encodes a group of transcription factors whose evolution has accompanied the major morphological and functional innovations of vertebrate species. The evolutionary conservation throughout diverse lineages of metazoan and the functional importance in development rendered Pax family an ideal system to address the relationship inside Chordata phylum. In the present study, we sequenced and annotated four genomic regions containing Chinese amphioxus (Branchiostoma belcheri) Pax genes, and retrieved homologous sequences from public database. In comparison with vertebrate homologues, the predicted amphioxus Pax proteins display high sequence conservation. Evidences from the molecular phylogenetic studies and gene organization analyses supports cephalochordates have a much closer relationship to vertebrates than that between tunicates and vertebrates, contrasting to urochordate relatives hypothesis proposed by several latest studies. Analysis of phylogenetic topology derived from concatenated subfamily datasets uncovered a potential statistical bias of supermatrix approach. Furthermore, we deduced an evolutionary scenario of Pax gene family. This scenario provided a plausible explanation for the origin and dynamics of the Pax gene members.

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The scallop is an economically important sea food prized for its large and delicious adductor muscle. Studying the molecular basis of scallop muscle growth is important for both scallop breeding and our understanding of muscle mass regulation in bivalve. Myostatin (MSTN) is a conserved negative regulator of muscle growth and development. Here we report the MSTN gene from Zhikong scallop (Chlamys farreri Jones et Preston 1904). The C. farreri MSTN consists of 11651 nucleotides encoding 457 amino acids. The gene has a 3-exon/2-intron structure that is conserved with vertebrate homologs. The exons are 586, 380 and 408 bp in length, respectively, and separated by introns of 5086 and 1518 bp. The protein sequence contains characteristic conserved residues including a cleavage motif of proteolysis (RXXR) and nine cysteines. Three transcription initiation sites were found at 62, 146, and 296 bp upstream of the translation start codon ATG. In silico analysis of the promoter region identified a TATA-box and several muscle-specific regulatory elements including COMP, MEF2s, MTBFs and E-boxes. Minisatellite DNA was found in intron 1. By fluorescence in situ hybridization (FISH), the gene was mapped to the long arm of a pair of middle subtelocentric chromosome. Quantitative analysis of MSTN transcripts in embryos/larvae indicated high expression level in gastrulae and limited expression at other stages. In adult scallops, MSTN is predominantly expressed in striated muscle, with different expression levels in other tissues. Our data provide valuable genomic and expression information which will aid the further study on scallop MSTN function and MSTN evolution.

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We determined the complete nucleotide sequences of mitochondrial (mt) genomes from two dicroglossid frogs, Hoplobatrachus tigerinus (Indian Bullfrog) and Euphlyctis hexadactylus (Indian Green frog). The genome sizes are 20462 bp in H. tigerinus and 20280 bp in E. hexadactylus. Although both genomes encode the typical 37 mt genes, the following unique features are observed: 1) the ND5 genes are duplicated in H. tigerinus that have completely identical sequences, whereas duplicated ND5 genes in E. hexadactylus possessed dissimilar substitutions; 2) duplicated control region (CR) in H. tigerinus has almost identical sequences whereas single control region (CR) was found in E. hexadactylus; 3) the tRNA-Leu (CUN) gene is translocated from the LTPF tRNA cluster to downstream of ND5-1 in H. tigerinus, and the tRNA-Pro gene is translocated from the LTPF tRNA cluster to downstream of CR in E. hexadactylus; 4) pseudo tRNA-Leu (CUN) and tRNA-Pro genes are observed in E. hexadactylus; and 5) two tRNA-Met genes are encoded in both species, as observed in the previously reported dicroglossid mt genomes. Almost all observed gene rearrangements in H. tigerinus and E. hexadactylus can be explained by the tandem duplication and random loss model, except translocation of tRNA-Pro in E. hexadactylus. The novel mt genomic features found in this study may be useful for future phylogenetic studies in the dicroglossid taxa. However, the mt genome with interesting features found in the present study reveal a high level of variation of gene order and gene content, inspiring more research to understand the mechanisms behind gene and genome evolution in the dicroglossid and as well as in the amphibian taxa in future studies.

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Six1, which belongs to the sine oculis homeobox (Six) protein family, is an evolutionarily conserved transcription factor found in diverse organisms ranging from flatworms to humans. Six1 is expressed in various tissues including the nervous system during ontogenesis and has been implicated in cell differentiation, morphogenesis, and organogenesis of the ganglia and sensory placodes. However, the molecular mechanisms by which Six1 influences these events at the transcriptional level remain largely unknown. In this study, we used ChIP-Display to discover genomic regions occupied in vivo by Six1 homeoprotein in the developing mouse embryo. To validate Six1 occupancy at each of Six1-bound regions, ChIP - Quantitative PCR was performed using locus-specific primers, and it showed robust enrichment of the Six1-bound sequences. To address their regulatory potential, each of the Six1-bound sequences was cloned into a reporter cassette containing beta-globin minimal promoter and lacZ gene and assayed for enhancer activity in transgenic mouse embryos. One of the novel sequences, which was designated Six1-bound Regulatory Element 1 (SRE1), was sufficient to activate lacZ reporter expression in the cranial and spinal ganglia. Comparative genomic analysis identified SRE1 sequences from a number of vertebrate phyla. Transgenic embryos carrying SRE1 sequences from human, chicken and frog showed reporter expression in a pattern similar to that of mouse SRE1, indicating their functional conservation. Through mutational analysis, we further showed that a conserved binding site matching the consensus for Six1/2/4/5 is required for the SRE1 regulatory activity. These data suggest that SRE1 is a functionally conserved transcriptional enhancer regulated by Six1.

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