Publications by authors named "Yasuko Akiyama-Oda"

25 Publications

  • Page 1 of 1

Hedgehog signaling controls segmentation dynamics and diversity via in a spider embryo.

Sci Adv 2020 Sep 9;6(37). Epub 2020 Sep 9.

Laboratory of Evolutionary Cell and Developmental Biology, JT Biohistory Research Hall, Takatsuki, Osaka, Japan.

Hedgehog (Hh) signaling plays fundamental roles in animal body patterning. Understanding its mechanistic complexity requires simple tractable systems that can be used for these studies. In the early spider embryo, Hh signaling mediates the formation of overall anterior-posterior polarity, yet it remains unclear what mechanisms link the initial Hh signaling activity with body axis segmentation, in which distinct periodic stripe-forming dynamics occur depending on the body region. We performed genome-wide searches for genes that transcriptionally respond to altered states of Hh signaling. Characterization of genes negatively regulated by Hh signaling suggested that , encoding a conserved transcription factor, functions as a key segmentation gene. Knockdown of prevented all dynamic processes causing spatial repetition of stripes, including temporally repetitive oscillations and bi-splitting, and temporally nonrepetitive tri-splitting. Thus, Hh signaling controls segmentation dynamics and diversity via These genome-wide data from an invertebrate illuminate novel mechanistic features of Hh-based patterning.
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http://dx.doi.org/10.1126/sciadv.aba7261DOI Listing
September 2020

Dataset on gene expressions affected by simultaneous knockdown of Hedgehog and Dpp signaling components in embryos of the spider .

Data Brief 2020 Feb 3;28:105088. Epub 2020 Jan 3.

Laboratory of Evolutionary Cell and Developmental Biology, JT Biohistory Research Hall, 1-1 Murasaki-cho, Takatsuki, Osaka, 569-1125, Japan.

Simultaneous, parental RNA interference (pRNAi) mediated knockdown of Hedgehog and Decapentaplegic (Dpp) signaling components, () and , respectively, exhibited serious defects in the formation of the major embryonic axes in the model spider . This paper describes a dataset of a custom oligonucleotide two-color microarray analysis that was carried out to compare the transcript expression levels between untreated (normal) and  +  double pRNAi embryos at late stage 5. Array spots that showed the intensity ratio of [ +  double pRNAi]/[normal] <0.6 were categorized as positive. The expressions of most, not all, of the transcripts related to the positive array spots were examined in embryos by whole-mount hybridization. Some of the stained embryos showed distinct patterns of gene expression. The data presented may be useful for characterizing the mechanisms of embryonic patterning in spider embryos.
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http://dx.doi.org/10.1016/j.dib.2019.105088DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7096682PMC
February 2020

The common house spider .

Evodevo 2020 20;11. Epub 2020 Mar 20.

1Laboratory of Evolutionary Cell and Developmental Biology, JT Biohistory Research Hall, 1-1 Murasaki-cho, Takatsuki, Osaka 569-1125 Japan.

The common house spider belonging to the Chelicerata in the phylum Arthropoda, has emerged as an experimental system for studying mechanisms of development from an evolutionary standpoint. In this article, we review the distinct characteristics of , the major research questions relevant to this organism, and the available key methods and resources. has a relatively short lifecycle and, once mated, periodically lays eggs. The morphogenetic field of the embryo is cellular from an early stage and exhibits stepwise symmetry-breaking events and stripe-forming processes that are associated with body axes formation and segmentation, respectively, before reaching the arthropod phylotypic stage. Self-regulatory capabilities of the embryonic field are a prominent feature in . The mechanisms and logic underlying the evolvability of heritable patterning systems at the phylum level could be one of the major avenues of research investigated using this animal. The sequenced genome reveals whole genome duplication (WGD) within chelicerates, which offers an invertebrate platform for investigating the potential roles of WGD in animal diversification and evolution. The development and evolution of lineage-specific organs, including the book lungs and the union of spinnerets and silk glands, are attractive subjects of study. Studies using can benefit from the use of parental RNA interference, microinjection applications (including cell labeling and embryonic RNA interference), multicolor fluorescence in situ hybridization, and laser ablation as well as rich genomic and transcriptomic resources. These techniques enable functional gene discoveries and the uncovering of cellular and molecular insights.
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http://dx.doi.org/10.1186/s13227-020-00152-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7082966PMC
March 2020

Microarray data on the comparison of transcript expression between normal and RNAi embryos in the common house spider .

Data Brief 2019 Aug 5;25:104350. Epub 2019 Aug 5.

Laboratory of Evolutionary Cell and Developmental Biology, JT Biohistory Research Hall, 1-1 Murasaki-cho, Takatsuki, Osaka 569-1125, Japan.

We conducted a custom microarray experiment to detect the differences in the transcript expression levels between untreated (normal) and -RNAi embryos at late stage 6 in the common house spider . The array probes were designed based on accumulated EST and cDNA sequences. The microarray dataset has been deposited in the Gene Expression Omnibus (GEO) Database at the National Center for Biotechnology Information (NCBI) under the accession GSE113064. The expression of the transcripts selected based on the detected differences was examined in embryos by whole-mount hybridization.
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http://dx.doi.org/10.1016/j.dib.2019.104350DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6702388PMC
August 2019

Experimental duplication of bilaterian body axes in spider embryos: Holm's organizer and self-regulation of embryonic fields.

Dev Genes Evol 2020 03 10;230(2):49-63. Epub 2019 Apr 10.

Laboratory of Evolutionary Cell and Developmental Biology, JT Biohistory Research Hall, 1-1 Murasaki-cho, Takatsuki, Osaka, 569-1125, Japan.

Bilaterally symmetric body plans of vertebrates and arthropods are defined by a single set of two orthogonal axes, the anterior-posterior (or head-tail) and dorsal-ventral axes. In vertebrates, and especially amphibians, complete or partial doubling of the bilaterian body axes can be induced by two different types of embryological manipulations: transplantation of an organizer region or bi-sectioning of an embryo. Such axis doubling relies on the ability of embryonic fields to flexibly respond to the situation and self-regulate toward forming a whole body. This phenomenon has facilitated experimental efforts to investigate the mechanisms of vertebrate body axes formation. However, few studies have addressed the self-regulatory capabilities of embryonic fields associated with body axes formation in non-vertebrate bilaterians. The pioneer spider embryologist Åke Holm reported twinning of spider embryos induced by both types of embryological manipulations in 1952; yet, his experiments have not been replicated by other investigators, and access to spider or non-vertebrate twins has been limited. In this review, we provide a historical background on twinning experiments in spiders, and an overview of current twinning approaches in familiar spider species and related molecular studies. Moreover, we discuss the benefits of the spider model system for a deeper understanding of the ancestral mechanisms of body axes formation in arthropods, as well as in bilaterians.
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http://dx.doi.org/10.1007/s00427-019-00631-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7128006PMC
March 2020

Homeobox Gene Duplication and Divergence in Arachnids.

Mol Biol Evol 2018 09;35(9):2240-2253

Department of Biological and Medical Sciences, Oxford Brookes University, Oxford, United Kingdom.

Homeobox genes are key toolkit genes that regulate the development of metazoans and changes in their regulation and copy number have contributed to the evolution of phenotypic diversity. We recently identified a whole genome duplication (WGD) event that occurred in an ancestor of spiders and scorpions (Arachnopulmonata), and that many homeobox genes, including two Hox clusters, appear to have been retained in arachnopulmonates. To better understand the consequences of this ancient WGD and the evolution of arachnid homeobox genes, we have characterized and compared the homeobox repertoires in a range of arachnids. We found that many families and clusters of these genes are duplicated in all studied arachnopulmonates (Parasteatoda tepidariorum, Pholcus phalangioides, Centruroides sculpturatus, and Mesobuthus martensii) compared with nonarachnopulmonate arachnids (Phalangium opilio, Neobisium carcinoides, Hesperochernes sp., and Ixodes scapularis). To assess divergence in the roles of homeobox ohnologs, we analyzed the expression of P. tepidariorum homeobox genes during embryogenesis and found pervasive changes in the level and timing of their expression. Furthermore, we compared the spatial expression of a subset of P. tepidariorum ohnologs with their single copy orthologs in P. opilio embryos. We found evidence for likely subfunctionlization and neofunctionalization of these genes in the spider. Overall our results show a high level of retention of homeobox genes in spiders and scorpions post-WGD, which is likely to have made a major contribution to their developmental evolution and diversification through pervasive subfunctionlization and neofunctionalization, and paralleling the outcomes of WGD in vertebrates.
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http://dx.doi.org/10.1093/molbev/msy125DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6107062PMC
September 2018

Genome-scale embryonic developmental profile of gene expression in the common house spider .

Data Brief 2018 Aug 24;19:865-867. Epub 2018 May 24.

Laboratory of Evolutionary Cell and Developmental Biology, JT Biohistory Research Hall, 1-1 Murasaki-cho, Takatsuki, Osaka 569-1125, Japan.

We performed RNA sequencing (RNA-Seq) at ten successive developmental stages in embryos of the common house spider . Two independent datasets from two pairs of parents represent the normalized coverage of mapped RNA-Seq reads along scaffolds of the genome assembly. Transcript abundance was calculated against existing AUGUSTUS gene models. The datasets have been deposited in the Gene Expression Omnibus (GEO) Database at the National Center for Biotechnology Information (NCBI) under the accession number GSE112712.
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http://dx.doi.org/10.1016/j.dib.2018.05.106DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5997937PMC
August 2018

A quantitative study of the diversity of stripe-forming processes in an arthropod cell-based field undergoing axis formation and growth.

Dev Biol 2018 05 16;437(2):84-104. Epub 2018 Mar 16.

Laboratory of Evolutionary Cell and Developmental Biology, JT Biohistory Research Hall, 1-1 Murasaki-cho, Takatsuki, Osaka 569-1125, Japan; Laboratory of Biohistory, Department of Biological Sciences, Graduate School of Science, Osaka University, Japan. Electronic address:

One of the conserved traits of arthropod embryonic development is striped expression of homologs of Drosophila segment polarity genes, including hedgehog (hh). Although a diversity of stripe-forming processes is recognized among arthropod embryos, such varied stripe-forming processes have not been well characterized from cellular and quantitative perspectives. The spider Parasteatoda tepidariorum embryo, which has a hh-dependent mechanism of axis formation, offers a cell-based field where the stripes of Pt-hh (a hh homolog) expression dynamically develop in accordance with axis formation and growth, with the patterning processes varying among the regions of the field. In this study, using cell labeling, we mapped the future body subdivisions to the germ disc in the spider embryo and provided substantial evidence for the occurrence of kinetic waves of Pt-hh expression in the presumptive head and opisthosomal (or abdominal) regions of the embryonic field. Notably, combined with cell tracking, we showed that surface cells at and near the center of the germ disc persist in the posterior portion of the field from where Pt-hh stripes sequentially arise, suggesting the operation of ordered oscillations of Pt-hh expression. We then conducted a quantitative analysis of forming/formed Pt-hh stripes using serially timed fixation of sibling embryos. By utilizing length measurements that reflect the axis growth of the embryonic field, we reconstructed the pattern dynamics, which captured repeated splitting of Pt-hh stripes and oscillations of Pt-hh expression in the presumptive head and opisthosomal regions, respectively. In the intermediate thoracic region, three stripes of Pt-hh expression showed a late appearance, with the segmental units specified much earlier by another mechanism. Analyses provided quantitative estimates related to axis growth and stripe-splitting and oscillation events, including the periods of the patterning cycles. This work characterizes the diversity of stripe-forming processes in a cell-based field in a common spatiotemporal framework and highlights the contrasting dynamics of splitting versus oscillation. The cellular and quantitative data presented here provide the foundation for experimental, theoretical and evolutionary studies of cell-based pattern formation, especially body axis segmentation in arthropods.
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http://dx.doi.org/10.1016/j.ydbio.2018.03.001DOI Listing
May 2018

The house spider genome reveals an ancient whole-genome duplication during arachnid evolution.

BMC Biol 2017 07 31;15(1):62. Epub 2017 Jul 31.

Human Genome Sequencing Center, Department of Molecular and Human Genetics, Baylor College of Medicine, One Baylor Plaza, Houston, TX, 77030, USA.

Background: The duplication of genes can occur through various mechanisms and is thought to make a major contribution to the evolutionary diversification of organisms. There is increasing evidence for a large-scale duplication of genes in some chelicerate lineages including two rounds of whole genome duplication (WGD) in horseshoe crabs. To investigate this further, we sequenced and analyzed the genome of the common house spider Parasteatoda tepidariorum.

Results: We found pervasive duplication of both coding and non-coding genes in this spider, including two clusters of Hox genes. Analysis of synteny conservation across the P. tepidariorum genome suggests that there has been an ancient WGD in spiders. Comparison with the genomes of other chelicerates, including that of the newly sequenced bark scorpion Centruroides sculpturatus, suggests that this event occurred in the common ancestor of spiders and scorpions, and is probably independent of the WGDs in horseshoe crabs. Furthermore, characterization of the sequence and expression of the Hox paralogs in P. tepidariorum suggests that many have been subject to neo-functionalization and/or sub-functionalization since their duplication.

Conclusions: Our results reveal that spiders and scorpions are likely the descendants of a polyploid ancestor that lived more than 450 MYA. Given the extensive morphological diversity and ecological adaptations found among these animals, rivaling those of vertebrates, our study of the ancient WGD event in Arachnopulmonata provides a new comparative platform to explore common and divergent evolutionary outcomes of polyploidization events across eukaryotes.
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http://dx.doi.org/10.1186/s12915-017-0399-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5535294PMC
July 2017

Evolutionary origin of type IV classical cadherins in arthropods.

BMC Evol Biol 2017 06 17;17(1):142. Epub 2017 Jun 17.

Laboratory of Evolutionary Cell and Developmental Biology, JT Biohistory Research Hall, 1-1 Murasaki-cho, Takatsuki, 569-1125, Osaka, Japan.

Background: Classical cadherins are a metazoan-specific family of homophilic cell-cell adhesion molecules that regulate morphogenesis. Type I and type IV cadherins in this family function at adherens junctions in the major epithelial tissues of vertebrates and insects, respectively, but they have distinct, relatively simple domain organizations that are thought to have evolved by independent reductive changes from an ancestral type III cadherin, which is larger than derived paralogs and has a complicated domain organization. Although both type III and type IV cadherins have been identified in hexapods and branchiopods, the process by which the type IV cadherin evolved is still largely unclear.

Results: Through an analysis of arthropod genome sequences, we found that the only classical cadherin encoded in chelicerate genomes was the type III cadherin and that the two type III cadherin genes found in the spider Parasteatoda tepidariorum genome exhibited a complex yet ancestral exon-intron organization in arthropods. Genomic and transcriptomic data from branchiopod, copepod, isopod, amphipod, and decapod crustaceans led us to redefine the type IV cadherin category, which we separated into type IVa and type IVb, which displayed a similar domain organization, except type IVb cadherins have a larger number of extracellular cadherin (EC) domains than do type IVa cadherins (nine versus seven). We also showed that type IVa cadherin genes occurred in the hexapod, branchiopod, and copepod genomes whereas only type IVb cadherin genes were present in malacostracans. Furthermore, comparative characterization of the type IVb cadherins suggested that the presence of two extra EC domains in their N-terminal regions represented primitive characteristics. In addition, we identified an evolutionary loss of two highly conserved cysteine residues among the type IVa cadherins of insects.

Conclusions: We provide a genomic perspective of the evolution of classical cadherins among bilaterians, with a focus on the Arthropoda, and suggest that following the divergence of early arthropods, the precursor of the insect type IV cadherin evolved through stepwise reductive changes from the ancestral type III state. In addition, the complementary distributions of polarized genomic characters related to type IVa/IVb cadherins may have implications for our interpretations of pancrustacean phylogeny.
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http://dx.doi.org/10.1186/s12862-017-0991-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5473995PMC
June 2017

Multi-color FISH facilitates analysis of cell-type diversification and developmental gene regulation in the Parasteatoda spider embryo.

Dev Growth Differ 2016 Feb 9;58(2):215-24. Epub 2016 Feb 9.

JT Biohistory Research Hall, 1-1 Murasaki-cho, 569-1125, Takatsuki, Osaka, 569-1125, Japan.

The simultaneous and quantitative analysis of the expression of multiple genes helps to shed light on gene regulatory networks. We established a method for multi-color fluorescence in situ hybridization (mFISH) for the analysis of cell-type diversification and developmental gene regulation in the embryo of the spider Parasteatoda tepidariorum. This mFISH technique allowed quadruple staining using four types of labels for RNA probes, digoxigenin, fluorescein, biotin, and dinitrophenyl, together with different fluorescent tyramides. To validate the usability of mFISH, we conducted four experiments. First, we distinguished similar gene expression patterns with mFISH, which showed overlaps and differences in the expression domains of anterior patterning hedgehog (hh), orthodenticle (otd), and labial genes at a cellular resolution. Second, we used mFISH to identify early cell types that are internalized on the anterior side. We found that fork head-positive cells were subdivided into two cell types, 012_A08-positive endoderm cells and twist-positive mesoderm cells. Third, we quantified the ratio of expression levels of the odd-paired (opa) gene in the chelicera and pedipalp segments based on the intensity of mFISH signals. Finally, we combined mFISH with embryonic RNA interference. It was possible to identify opa knockdown cell clones and detect the specific reduction of opa and the upregulation of otd and hh expression levels in the same cell clone that formed in the head region. This study proposes that mFISH is a powerful tool for the cell-level analysis of gene regulation and quantification in the spider model.
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http://dx.doi.org/10.1111/dgd.12263DOI Listing
February 2016

New directions of EvoDevo: revisiting ideas of professor Hotta.

J Neurogenet 2012 Mar 12;26(1):25-7. Epub 2012 Jan 12.

JT Biohistory Research Hall, Osaka, Japan.

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http://dx.doi.org/10.3109/01677063.2011.642429DOI Listing
March 2012

Travelling and splitting of a wave of hedgehog expression involved in spider-head segmentation.

Nat Commun 2011 Oct 11;2:500. Epub 2011 Oct 11.

JT Biohistory Research Hall, Murasaki-cho, Takatsuki, Osaka, Japan.

During development segmentation is a process that generates a spatial periodic pattern. Peak splitting of waves of gene expression is a mathematically predicted, simple strategy accounting for this type of process, but it has not been well characterized biologically. Here we show temporally repeated splitting of gene expression into stripes that is associated with head axis growth in the spider Achaearanea embryo. Preceding segmentation, a wave of hedgehog homologue gene expression is observed to travel posteriorly during development stage 6. This stripe, co-expressing an orthodenticle homologue, undergoes two cycles of splitting and shifting accompanied by convergent extension, serving as a generative zone for the head segments. The two orthodenticle and odd-paired homologues are identified as targets of Hedgehog signalling, and evidence suggests that their activities mediate feedback to maintain the head generative zone and to promote stripe splitting in this zone. We propose that the 'stripe-splitting' strategy employs genetic components shared with Drosophila blastoderm subdivision, which are required for participation in an autoregulatory signalling network.
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http://dx.doi.org/10.1038/ncomms1510DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3207210PMC
October 2011

Early embryonic development in the spider Achaearanea tepidariorum: Microinjection verifies that cellularization is complete before the blastoderm stage.

Arthropod Struct Dev 2010 Nov;39(6):436-45

JT Biohistory Research Hall, 1-1 Murasaki-cho, Takatsuki, Osaka, Japan.

The spider Achaearanea tepidariorum is emerging as a non-insect model for studying developmental biology. However, the availability of microinjection into early embryos of this spider has not been reported. We defined the early embryonic stages in A. tepidariorum and applied microinjection to its embryos. During the preblastoderm 16- and 32-nucleus stages, the energids were moving toward the egg periphery. When fluorochrome-conjugated dextran was microinjected into the peripheral region of 16-nucleus stage embryos, it was often incorporated into a single energid and inherited in the progeny without leaking out to surrounding energids. This suggested that 16-nucleus stage embryos consisted of compartments, each containing a single energid. These compartments were considered to be separate cells. Fluorochrome-conjugated dextran could be introduced into single cells of 16- to 128-nucleus stage embryos, allowing us to track cell fate and movement. Injection with mRNA encoding a nuclear localization signal/green fluorescent protein fusion construct demonstrated exogenous expression of the protein in live spider embryos. We propose that use of microinjection will facilitate studies of spider development. Furthermore, these data imply that in contrast to the Drosophila syncytial blastoderm embryo, the cell-based structure of the Achaearanea blastoderm embryo restricts diffusion of cytoplasmic gene products.
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http://dx.doi.org/10.1016/j.asd.2010.05.009DOI Listing
November 2010

Cell migration that orients the dorsoventral axis is coordinated with anteroposterior patterning mediated by Hedgehog signaling in the early spider embryo.

Development 2010 Apr;137(8):1263-73

JT Biohistory Research Hall, 1-1 Murasaki-cho, Takatsuki, Osaka, Japan.

The early embryo of the spider Achaearanea tepidariorum is emerging as a model for the simultaneous study of cell migration and pattern formation. A cell cluster internalized at the center of the radially symmetric germ disc expresses the evolutionarily conserved dorsal signal Decapentaplegic. This cell cluster migrates away from the germ disc center along the basal side of the epithelium to the germ disc rim. This cell migration is thought to be the symmetry-breaking event that establishes the orientation of the dorsoventral axis. In this study, knockdown of a patched homolog, At-ptc, that encodes a putative negative regulator of Hedgehog (Hh) signaling, prevented initiation of the symmetry-breaking cell migration. Knockdown of a smoothened homolog, At-smo, showed that Hh signaling inactivation also arrested the cells at the germ disc center, whereas moderate inactivation resulted in sporadic failure of cell migration termination at the germ disc rim. hh transcript expression patterns indicated that the rim and outside of the germ disc were the source of the Hh ligand. Analyses of patterning events suggested that in the germ disc, short-range Hh signal promotes anterior specification and long-range Hh signal represses caudal specification. Moreover, negative regulation of Hh signaling by At-ptc appears to be required for progressive derepression of caudal specification from the germ disc center. Cell migration defects caused by At-ptc and At-smo knockdown correlated with patterning defects in the germ disc epithelium. We propose that the cell migration crucial for dorsoventral axis orientation in Achaearanea is coordinated with anteroposterior patterning mediated by Hh signaling.
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http://dx.doi.org/10.1242/dev.045625DOI Listing
April 2010

Differing strategies for forming the arthropod body plan: lessons from Dpp, Sog and Delta in the fly Drosophila and spider Achaearanea.

Dev Growth Differ 2008 May;50(4):203-14

JT Biohistory Research Hall, 1-1 Murasaki-cho, Takatsuki, Osaka 569-1125, Japan.

In the insect Drosophila embryo, establishment of maternal transcription factor gradients, rather than cell-cell interactions, is fundamental to patterning the embryonic axes. In contrast, in the chelicerate spider embryo, cell-cell interactions are thought to play a crucial role in the development of the embryonic axes. A grafting experiment by Holm using spider eggs resulted in duplication of the embryonic axes, similar to the Spemann's organizer experiment using amphibian eggs. Recent work using the house spider Achaearanea tepidariorum has demonstrated that the homologs of decapentaplegic (dpp), short gastrulation (sog) and Delta, which encode a bone morphogenetic protein (BMP)-type ligand, its antagonist and a Notch ligand, respectively, are required in distinct aspects of axis formation. Achaearanea Dpp appears to function as a symmetry-breaking signal, which could account for Holm's results to some extent. Experimental findings concerning Achaearanea sog and Delta have highlighted differences in the mechanisms underlying ventral and posterior development between Drosophila and Achaearanea. Achaearanea ventral patterning essentially depends on sog function, in contrast to the Drosophila patterning mechanism, which is based on the nuclear gradient of Dorsal. Achaearanea posterior (or opisthosomal) patterning relies on the function of the caudal lobe, which develops from cells surrounding the blastopore through progressive activation of Delta-Notch signaling. In this review, we describe the differing strategies for forming the arthropod body plan in the fly and spider, and provide a perspective towards understanding the relationship between the arthropod and vertebrate body plans.
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http://dx.doi.org/10.1111/j.1440-169X.2008.00998.xDOI Listing
May 2008

Progressive activation of Delta-Notch signaling from around the blastopore is required to set up a functional caudal lobe in the spider Achaearanea tepidariorum.

Development 2007 Jun 16;134(12):2195-205. Epub 2007 May 16.

JT Biohistory Research Hall, 1-1 Murasaki-cho, Takatsuki, Osaka 569-1125, Japan.

In the development of most arthropods, the caudal region of the elongating germ band (the growth zone) sequentially produces new segments. Previous work with the spider Cupiennius salei suggested involvement of Delta-Notch signaling in segmentation. Here, we report that, in the spider Achaearanea tepidariorum, the same signaling pathway exerts a different function in the presumptive caudal region before initiation of segmentation. In the developing spider embryo, the growth zone becomes morphologically apparent as a caudal lobe around the closed blastopore. We found that, preceding caudal lobe formation, transcripts of a Delta homolog, At-Delta, are expressed in evenly spaced cells in a small area covering the closing blastopore and then in a progressively wider area of the germ disc epithelium. Cells with high At-Delta expression are likely to be prospective mesoderm cells, which later express a twist homolog, At-twist, and individually internalize. Cells remaining at the surface begin to express a caudal homolog, At-caudal, to differentiate as caudal ectoderm. Knockdown of At-Delta by parental RNA interference results in overproduction of At-twist-expressing mesoderm cells at the expense of At-caudal-expressing ectoderm cells. This condition gives rise to a disorganized caudal region that fails to pattern the opisthosoma. In addition, knockdown of Notch and Suppressor of Hairless homologs produces similar phenotypes. We suggest that, in the spider, progressive activation of Delta-Notch signaling from around the blastopore leads to stochastic cell fate decisions between mesoderm and caudal ectoderm through a process of lateral inhibition to set up a functional caudal lobe.
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http://dx.doi.org/10.1242/dev.004598DOI Listing
June 2007

Axis specification in the spider embryo: dpp is required for radial-to-axial symmetry transformation and sog for ventral patterning.

Development 2006 Jun;133(12):2347-57

JT Biohistory Research Hall, 1-1 Murasaki-cho, Takatsuki, Osaka 569-1125, Japan.

The mechanism by which Decapentaplegic (Dpp) and its antagonist Short gastrulation (Sog) specify the dorsoventral pattern in Drosophila embryos has been proposed to have a common origin with the mechanism that organizes the body axis in the vertebrate embryo. However, Drosophila Sog makes only minor contributions to the development of ventral structures that hypothetically correspond to the vertebrate dorsum where the axial notochord forms. In this study, we isolated a homologue of the Drosophila sog gene in the spider Achaearanea tepidariorum, and characterized its expression and function. Expression of sog mRNA initially appeared in a radially symmetrical pattern and later became confined to the ventral midline area, which runs axially through the germ band. RNA interference-mediated depletion of the spider sog gene led to a nearly complete loss of ventral structures, including the axial ventral midline and the central nervous system. This defect appeared to be the consequence of dorsalization of the ventral region of the germ band. By contrast, the extra-embryonic area formed normally. Furthermore, we showed that embryos depleted for a spider homologue of dpp failed to break the radial symmetry, displaying evenly high levels of sog expression except in the posterior terminal area. These results suggest that dpp is required for radial-to-axial symmetry transformation of the spider embryo and sog is required for ventral patterning. We propose that the mechanism of spider ventral specification largely differs from that of the fly. Interestingly, ventral specification in the spider is similar to the process in vertebrates in which the antagonism of Dpp/BMP signaling plays a central role in dorsal specification.
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http://dx.doi.org/10.1242/dev.02400DOI Listing
June 2006

Diversification of epithelial adherens junctions with independent reductive changes in cadherin form: identification of potential molecular synapomorphies among bilaterians.

Evol Dev 2005 Sep-Oct;7(5):376-89

JT Biohistory Research Hall, 1-1 Murasaki-cho, Takatsuki, Osaka 569-1125, Japan.

The adherens junction (AJ) is the most universal junction found in bilaterian epithelia and may represent one of the earliest types of cell-cell junctions. The adhesion molecules responsible for forming AJs are the classic cadherins (referred to simply as cadherins), whose extracellular domain organization displays marked variety among species examined so far. In this study, we attempted to reconstruct the evolution of cadherin by analyzing new data from several arthropods (two insects, one non-insect hexapod, three crustaceans, and one chelicerate) and previously published sequences for Drosophila melanogaster and other animals. The results of comparative analyses using the BLAST tool and immunohistochemical analyses revealed that the extracellular domain organizations of a decapod, an isopod, a spider, and a starfish cadherin, which are present at AJs in the embryonic epithelia are homologous. Independent reductive changes from the ancestral state were evident in the epithelia of hexapods+branchiopod, vertebrates+urochordates, and a cephalochordate. The form of cadherins in hexapods is more closely related to that of a branchiopod than to that of malacostracan crustaceans, and one of those of vertebrates is more closely related to that of urochordates than to that of a cephalochordate. Although the sampling of taxa is limited at this stage of research, we hypothesize that the reductive events in cadherin structure related to AJ formation in the epithelia may possess information about bilaterian relationships as molecular synapomorphies.
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http://dx.doi.org/10.1111/j.1525-142X.2005.05043.xDOI Listing
December 2005

Expression patterns of a twist-related gene in embryos of the spider Achaearanea tepidariorum reveal divergent aspects of mesoderm development in the fly and spider.

Zoolog Sci 2005 Feb;22(2):177-85

JT Biohistory Research Hall, Osaka, Japan.

We cloned an Achaearanea tepidariorum (Chelicerata, Arachnida) gene related to Drosophila twist (twi), which encodes a basic helix-loop-helix transcription factor required to specify mesoderm fate in the Drosophila embryo. The cloned spider gene was designated At.twist (At.twi). We examined its expression by whole-mount in situ hybridization. At.twi transcripts were first detected in cells located at the polar and equatorial areas of the spherical embryo when the cumulus reached the equator. As the extra-embryonic area expanded, more cells expressed At.twi transcripts. The At.twi-expressing cells became distributed nearly uniformly in the embryonic area. At these stages, some At.twi-expressing cells were found in the surface epithelial cell layer, but other At.twi-expressing cells were at slightly deeper positions from the surface. When the embryo was transformed into a germ band, all At.twi-expressing cells were situated just beneath the surface ectoderm, where they became metamerically arranged. Although little expression was observed in the caudal lobe of the elongating germ band, new stripes of At.twi expression appeared beneath the ectoderm in accordance with the posterior growth. These observations suggested that the cells expressing At.twi were most likely mesoderm. We propose that At.twi can be used as a molecular marker for analyzing mesoderm development in the spider embryo. Moreover, comparison of the expression patterns of twi and At.twi revealed divergent aspects of mesoderm development in the fly and spider. In addition, we cloned an Achaearanea gene related to snail, which is another mesoderm-determining gene in Drosophila, and showed that its expression was restricted to the ectoderm with no indication for a role in mesoderm development.
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http://dx.doi.org/10.2108/zsj.22.177DOI Listing
February 2005

Two classic cadherin-related molecules with no cadherin extracellular repeats in the cephalochordate amphioxus: distinct adhesive specificities and possible involvement in the development of multicell-layered structures.

J Cell Sci 2004 Jun 18;117(Pt 13):2757-67. Epub 2004 May 18.

JT Biohistory Research Hall, 1-1 Murasaki-cho, Takatsuki, Osaka 569-1125, Japan.

We previously reported the existence of Bb-cadherin, a molecule related to classic cadherin, in the cephalochordate amphioxus (Branchiostoma belcheri). The structure of Bb-cadherin is unique in that it lacks the cadherin extracellular repeats, although its cytoplasmic domain shows close similarities to those of typical classic cadherins. The extracellular region of Bb-cadherin consists of laminin globular domains and a cysteine-rich EGF-like domain that are similar to domains in nonchordate classic cadherins. In this study, we identified a second amphioxus cadherin. It was designated Bb2-cadherin (Bb2C) while the previously reported cadherin has been renamed Bb1-cadherin (Bb1C). Bb2C is very similar to Bb1C in its overall structure and amino acid sequence. Genomic BLAST searches and phylogenetic analyses suggested that these two amphioxus genes have been generated through a gene duplication that occurred after separation of the cephalochordates from the other animals. They also bear distinct adhesive specificities. Immunohistochemical analyses showed that Bb1C and Bb2C, together with beta-catenin, appear to function as adherens junction constituents in the epithelia of different germ layers of the amphioxus embryo. Differential expression of the two cadherins was also observed in the developing, multicell-layered notochord. These observations suggest that, despite their unique structures, the functions and developmental roles of Bb1C and Bb2C are comparable to those of the classic cadherins characterized to date in other animal groups, such as the vertebrate E- and N-cadherins and the Drosophila DE- and DN-cadherins. The possible involvement of Bb1C and Bb2C in the development of multicell-layered structures characteristic of the cephalochordate body plan is presented.
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http://dx.doi.org/10.1242/jcs.01045DOI Listing
June 2004

NMR and ICP spectroscopic analysis of the DNA-binding domain of the Drosophila GCM protein reveals a novel Zn2+ -binding motif.

Protein Eng 2003 Apr;16(4):247-54

Biomolecular Engineering Research Institute, 6-2-3 Furue-dai, Suita, Osaka 560-0874, Japan.

Drosophila GCM (glial cell missing) is a novel DNA-binding protein that determines the fate of glial precursors from the neural default to glia. The GCM protein contains the functional domain that is essential for recognition of the upstream sequence of the repo gene. In the DNA-binding region of this GCM protein, there is a cysteine-rich region with which divalent metal ions such as Zn(2+) must bind and other proteins belonging to the GCM family have a corresponding region. To obtain a more detailed insight into the structural and functional features of this DNA-binding region, we have determined the minimal DNA-binding domain and obtained inductively coupled plasma atomic emission spectra and (1)H-(15)N, (1)H-(15)N-(13)C and (113)Cd(2+) NMR spectra, with or without its specific DNA molecule. Considering the results, it was concluded that the minimal DNA-binding domain includes two Zn(2+)-binding sites, one of which is adjacent to the interface for DNA binding. Systematic mutational analyses of the conserved cysteine residues in the minimal DNA-binding domain revealed that one Zn(2+)-binding site is indispensable for stabilization of the higher order structure of this DNA-binding domain, but that the other is not.
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http://dx.doi.org/10.1093/proeng/gzg040DOI Listing
April 2003

Early patterning of the spider embryo: a cluster of mesenchymal cells at the cumulus produces Dpp signals received by germ disc epithelial cells.

Development 2003 May;130(9):1735-47

JT Biohistory Research Hall, 1-1 Murasaki-cho, Takatsuki, Osaka 569-1125, Japan.

In early embryogenesis of spiders, the cumulus is characteristically observed as a cellular thickening that arises from the center of the germ disc and moves centrifugally. This cumulus movement breaks the radial symmetry of the germ disc morphology, correlating with the development of the dorsal region of the embryo. Classical experiments on spider embryos have shown that a cumulus has the capacity to induce a secondary axis when transplanted ectopically. In this study, we have examined the house spider, Achaearanea tepidariorum, on the basis of knowledge from Drosophila to characterize the cumulus at the cellular and molecular level. In the cumulus, a cluster of about 10 mesenchymal cells, designated the cumulus mesenchymal (CM) cells, is situated beneath the epithelium, where the CM cells migrate to the rim of the germ disc. Germ disc epithelial cells near the migrating CM cells extend cytoneme-like projections from their basal side onto the surface of the CM cells. Molecular cloning and whole-mount in situ hybridization showed that the CM cells expressed a spider homolog of Drosophila decapentaplegic (dpp), which encodes a secreted protein that functions as a dorsal morphogen in the Drosophila embryo. Furthermore, the spider Dpp signal appeared to induce graded levels of the phosphorylated Mothers against dpp (Mad) protein in the nuclei of germ disc epithelial cells. Adding data from spider homologs of fork head, orthodenticle and caudal, we suggest that, in contrast to the Drosophila embryo, the progressive mesenchymal-epithelial cell interactions involving the Dpp-Mad signaling cascade generate dorsoventral polarity in accordance with the anteroposterior axis formation in the spider embryo. Our findings support the idea that the cumulus plays a central role in the axial pattern formation of the spider embryo.
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http://dx.doi.org/10.1242/dev.00390DOI Listing
May 2003

A novel amphioxus cadherin that localizes to epithelial adherens junctions has an unusual domain organization with implications for chordate phylogeny.

Evol Dev 2002 Nov-Dec;4(6):426-34

Tsukita Cell Axis Project, ERATO, Japan Science and Technology Corporation, Yoshidakonoe-cho, Sakyo-ku, Kyoto 606-8501, Japan.

Although data are available from only vertebrates, urochordates, and three nonchordate animals, there are definite differences in the structures of classic cadherins between vertebrates plus urochordates and nonchordates. In this study we examined structural diversity of classic cadherins among bilaterian animals by obtaining new data from an amphioxus (Cephalochordata, Chordata), an acorn worm (Hemichordata), a sea star (Echinodermata), and an oyster (Mollusca). The structures of newly identified nonchordate cadherins are grouped together with those of the known sea urchin and Drosophila cadherins, whereas the structure of an amphioxus (Branchiostoma belcheri) cadherin, designated BbC, is differently categorized from those of other known chordate cadherins. BbC is identified as a cadherin by its cytoplasmic domain whose sequence is highly related to the cytoplasmic sequences of all known classic cadherins, but it lacks all of the five repeats constituting the extracellular homophilic-binding domain of other chordate cadherins. The ectodomains of BbC match the ectodomains found in nonchordate cadherins but not present in other chordate cadherins. We show that the BbC functions as a cell-cell adhesion molecule when expressed in Drosophila S2 cells and localizes to adherens junctions in the ectodermal epithelia in amphioxus embryos. We argue that BbC is the amphioxus homologue of the classic cadherins involved in the formation of epithelial adherens junctions. The structural relationships of the cadherin molecules allow us to propose a possibility that cephalochordates might be basal to the sister-groups vertebrates and urochordates.
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http://dx.doi.org/10.1046/j.1525-142x.2002.02031.xDOI Listing
May 2003