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Ciona Intestinalis (ciona + intestinali)
Kinds of Ciona Intestinalis Selected AbstractsMechanism of DNA replication-dependent transcriptional activation of the acetylcholinesterase gene in the Ciona intestinalis embryoDEVELOPMENT GROWTH & DIFFERENTIATION, Issue 9 2009Yumiko Kataoka The acetylcholinesterase-encoding gene in the ascidian Ciona intestinalis (Ci-AChE) is expressed in tail muscle cells from the gastrula stage. When the embryo was continuously treated with aphidicolin from the 32-cell stage, Ci-AChE was not expressed even when control embryos reached the tailbud stage. This result suggests that Ci-AChE acquires the competence to be transcribed after passing through a certain number of DNA replication cycles. A lacZ reporter gene containing the 5, flanking region of Ci-AChE was expressed in the tail muscle cells. Aphidicolin treatment from the 32-cell stage affected, but did not completely suppress, the expression of lacZ. A bisulfite sequencing analysis was carried out to examine the methylation status of four regions within the 5, flanking sequence and the first exon. However, all of these regions remained unmethylated from the 16-cell to 110-cell stages. The results suggested that the DNA of the Ci-AChE locus is not responsible for counting the rounds of replication. We examined the expression of the C. intestinalis MyoD (Ci-MyoD), a transcription factor that activates Ci-AChE. Aphidicolin treatment from the 32-cell stage suppressed the expression of Ci-MyoD, even when control embryos reached the gastrula stage. These results suggest that a lack of Ci-MyoD is critical to the suppression of Ci-AChE in aphidicolin-treated embryos. [source] Novel genes involved in canonical Wnt/, -catenin signaling pathway in early Ciona intestinalis embryosDEVELOPMENT GROWTH & DIFFERENTIATION, Issue 4 2008Shuichi Wada We report here characterization of five genes for novel components of the canonical Wnt/, -catenin signaling pathway. These genes were identified in the ascidian Ciona intestinalis through a loss-of-function screening for genes required for embryogenesis with morpholinos, and four of them have counterparts in vertebrates. The five genes we studied are as follows: Ci-PGAP1, a Ciona orthologue of human PGAP1, which encodes GPI (glycosylphosphatidylinositol) inositol-deacylase, Ci-ZF278, a gene encoding a C2H2 zinc-finger protein, Ci-C10orf11, a Ciona orthologue of human C10orf11 that encodes a protein with leucine-rich repeats, Ci-Spatial/C4orf17, a single counterpart for two human genes Spatial and C4orf17, and Ci-FLJ10634, a Ciona orthologue of human FLJ10634 that encodes a member of the J-protein family. Knockdown of each of the genes mimicked , -catenin knockdown and resulted in suppression of the expression of , -catenin downstream genes (Ci-FoxD, Ci-Lhx3, Ci-Otx and Ci-Fgf9/16/20) and subsequent endoderm formation. For every gene, defects in knockdown embryos were rescued by overexpression of a constitutively active form, but not wild-type, of Ci- , -catenin. Dosage-sensitive interactions were found between Ci-,-catenin and each of the genes. These results suggest that these five genes act upstream of or parallel to Ci- , -catenin in the Wnt/, -catenin signaling pathway in early Ciona embryos. [source] Brachyury -downstream notochord genes and convergent extension in Ciona intestinalis embryosDEVELOPMENT GROWTH & DIFFERENTIATION, Issue 5 2007Kohji Hotta Formation of the chordate body is accomplished by a complex set of morphogenetic movements including convergent extension of notochord cells. In the ascidian Ciona intestinalis, Brachyury plays a key role in the formation of the notochord, and more than 30 Bra-downstream notochord genes have been identified. In the present study, we examined the effects of functional suppression of nine Bra -downstream notochord genes, which include Ci-PTP, Ci-ACL, Ci-prickle, Ci-netrin, Ci-trop, Ci-Noto3, Ci-ASAK, Ci-ERM and Ci-pellino. When the function of the first two genes (Ci-PTP and Ci-ACL) was suppressed with specific morpholinos, the notochord cells failed to converge, while functional suppression of Ci-prickle resulted in a failure of intercalation, and therefore the cells in these three types of embryo remained in the mid-dorsal region of the embryo. Functional suppression of the next four genes (Ci-netrin, Ci-trop, Ci-Noto3 and Ci-ASAK) resulted in the partial defect of intercalation, and the notochord did not consist of a single row. In addition, when the function of the last two genes (Ci-ERM and Ci-pellino) was suppressed, notochord cells failed to elongate in the embryo, even though convergence/extension took place normally. These results indicate that many Bra -downstream notochord genes are involved in convergence/extension of the embryo. [source] Network structure of projections extending from peripheral neurons in the tunic of ascidian larvaDEVELOPMENTAL DYNAMICS, Issue 8 2010Hiroshi Q. Terakubo Abstract In ascidian Ciona intestinalis, a subset of trunk epidermal neurons were shown to possess external network of neural projections. To characterize a more complete network in naturally hatched (chorionated) larvae, we visualized the structure with a confocal laser scanning microscope. High resolution images revealed the huge network consisting of several subnetworks in whole-larval tunic. We named this network the ASNET (ascidian dendritic network in tunic). The ASNET was dynamically generated and collapsed during larval stages. Interestingly, one of the subnetworks found around apical trunk epidermal neurons was bilaterally asymmetric. In caudal epidermal neurons, transmission electron microscopy revealed that 9+2 axonemes were accompanied by a vesicle-containing mass in the ASNET arbor, but the distal end of the arbor contained only the vesicle-containing fibrous mass and no 9+2 axonemes. The characteristics of the ASNET suggest that it forms a unique outer body network in the ascidian larval tunic. Developmental Dynamics 239:2278,2287, 2010.© 2010 Wiley-Liss, Inc. [source] Efficient transposition of a single Minos transposon copy in the genome of the ascidian Ciona intestinalis with a transgenic line expressing transposase in eggsDEVELOPMENTAL DYNAMICS, Issue 4 2010Akiko Hozumi Abstract Transgenesis with transposons is an important technique for studying genetic functions. In the ascidian Ciona intestinalis, methods for germline transformation with the Tc1/mariner transposon Minos have been established. A system to remobilize a single Minos copy in the genome is needed to refine this transgenic technique. In this study, such an experimental system was established with a transgenic line expressing Minos transposase in eggs. In the eggs of a double transgenic animal from a cross between the egg transposase line and a transgenic line having a single Minos insertion, the transposon was transposed into new positions of the Ciona genome, thus creating new insertions. Some of the new insertions caused enhancer detection. The majority of the new insertion sites were mapped on different chromosomes from that of the transposon donor. This characteristic of Minos is in contrast to that of the Sleeping Beauty transposon, which causes frequent intrachromosomal transposition. Developmental Dynamics 239:1076,1088, 2010. © 2010 Wiley-Liss, Inc. [source] Enhancer detection in the ascidian Ciona intestinalis with transposase-expressing lines of MinosDEVELOPMENTAL DYNAMICS, Issue 1 2008Yasunori Sasakura Abstract Germline transgenesis with a Tc1/mariner superfamily Minos transposon was achieved in the ascidian Ciona intestinalis. Transgenic lines that express transposases in germ cells are very useful for remobilizing transposon copies. In the present study, we created transposase-expressing lines of Minos in Ciona. A Ciona gene encoding protamine (Ci - prm) is expressed in the testes and sperm. Transgenic lines expressing Minos transposase in the testes and sperm were created with a cis -element of Ci - prm, and used for enhancer detection. Double-transgenic animals between transposase lines and a transgenic line with an enhancer detection vector passed on several independent enhancer detection events to subsequent progeny. This technique allowed us to isolate transgenic lines that express GFP in restricted tissues. This system provides an easy and efficient method for large-scale enhancer detection in Ciona intestinalis. Developmental Dynamics 237:39,50, 2008. © 2007 Wiley-Liss, Inc. [source] Postplasmic/PEM RNAs: A class of localized maternal mRNAs with multiple roles in cell polarity and development in ascidian embryosDEVELOPMENTAL DYNAMICS, Issue 7 2007François Prodon Abstract Ascidian is a good model to understand the cellular and molecular mechanisms responsible for mRNA localization with the discovery of a large family of localized maternal mRNAs, called postplasmic/PEM RNAs, which includes more than 40 members in three different ascidian species (Halocynthia roretzi, Ciona intestinalis, and C. savignyi). Among these mRNAs, two types (Type I and Type II) have been identified and show two different localization patterns from fertilization to the eight-cell stage. At the eight-cell stage, both types concentrate to a macromolecular cortical structure called CAB (for Centrosome Attracting Body) in the posterior-vegetal B4.1 blastomeres. The CAB is responsible for unequal cleavages and the partitioning of postplasmic/PEM RNAs at the posterior pole of embryos during cleavage stages. It has also been suggested that the CAB region could contain putative germ granules. In this review, we discuss recent data obtained on the distribution of Type I postplasmic/PEM RNAs from oogenesis to late development, in relation to their localization and translational control. We have first regrouped localization patterns for Type I and Type II into a comparative diagram and included all important definitions in the field. We also have made an exhaustive classification of their embryonic expression profiles (Type I or Type II), and analyzed their functions after knockdown and/or overexpression experiments and the role of the 3,-untranslated region (3,UTR) controlling both their localization and translation. Finally, we propose a speculative model integrating recent data, and we also discuss the relationship between postplasmic/PEM RNAs, posterior specification, and germ cell formation in ascidians. Developmental Dynamics 236:1698,1715, 2007. © 2007 Wiley-Liss, Inc. [source] Lineage-independent mosaic expression and regulation of the Ciona multidom gene in the ancestral notochordDEVELOPMENTAL DYNAMICS, Issue 7 2007Izumi Oda-Ishii Abstract The transcription factor Ciona Brachyury (Ci-Bra) plays an essential role in notochord development in the ascidian Ciona intestinalis. We characterized a putative Ci-Bra target gene, which we named Ci - multidom, and analyzed in detail its expression pattern in normal embryos and in embryos where Ci - Bra was misexpressed. Ci - multidom encodes a novel protein, which contains eight CCP domains and a partial VWFA domain. We show that an EGFP-multidom fusion protein localizes preferentially to the endoplasmic reticulum (ER), and is excluded from the nucleus. In situ hybridization experiments demonstrate that Ci - multidom is expressed in the notochord and in the anterior neural boundary (ANB). We found that the expression in the ANB is fully recapitulated by an enhancer element located upstream of Ci - multidom. By means of misexpression experiments, we provide evidence that Ci-Bra controls transcription of Ci - multidom in the notochord; however, while Ci-Bra is homogeneously expressed throughout this structure, Ci - multidom is transcribed at detectable levels only in a random subset of notochord cells. The number of notochord cells expressing Ci - multidom varies among different embryos and is independent of developmental stage, lineage, and position along the anterior,posterior axis. These results suggest that despite its morphological simplicity and invariant cell-lineage, the ancestral notochord is a mosaic of cells in which the gene cascade downstream of Brachyury is differentially modulated. Developmental Dynamics 236:1806,1819, 2007. © 2007 Wiley-Liss, Inc. [source] Novel genes involved in Ciona intestinalis embryogenesis: Characterization of gene knockdown embryosDEVELOPMENTAL DYNAMICS, Issue 7 2007Mayuko Hamada Abstract The sequenced genome of the urochordate ascidian Ciona intestinalis contains nearly 2,500 genes that have vertebrate homologues, but their functions are as yet unknown. To identify novel genes involved in early chordates embryogenesis, we previously screened 200 Ciona genes by knockdown experiments using specific morpholino oligonucleotides and found that suppression of the translation of 40 genes caused embryonic defects (Yamada et al. [2003] Development 130:6485,6495). We have since examined an additional 304 genes, that is, screening 504 genes overall, and a total of 111 genes showed morphological defects when gene function was suppressed. We further examined the role of these genes in the differentiation of six major tissues of the embryo: endoderm, muscle, epidermis, neural tissue, mesenchyme, and notochord. Based on the similarity of phenotypes of gene knockdown embryos, genes were categorized into several groups, with the suggestion that the genes within a given group are involved in similar developmental processes. For example, five were shown to be novel genes that are likely involved in ,-catenin,mediated endoderm formation. The type of large-scale screening used is, therefore, a powerful approach to identify novel genes with significant developmental functions, the details of which will be determined in future studies. Developmental Dynamics 236:1820,1831, 2007. © 2007 Wiley-Liss, Inc. [source] Optimized green fluorescent protein variants provide improved single cell resolution of transgene expression in ascidian embryosDEVELOPMENTAL DYNAMICS, Issue 2 2006Robert W. Zeller Abstract The green fluorescent protein (GFP) is used extensively to monitor gene expression and protein localization in living cells, particularly in developing embryos from a variety of species. Several GFP mutations have been characterized that improve protein expression and alter the emission spectra to produce proteins that emit green, blue, cyan, and yellow wavelengths. DsRed and its variants encode proteins that emit in the orange to red wavelengths. Many of these commercially available fluorescent proteins have been "codon optimized" for maximal levels of expression in mammalian cells. We have generated several fluorescent protein color variants that have been codon optimized for maximal expression in the ascidian Ciona intestinalis. By analyzing quantitative time-lapse recordings of transgenic embryos, we demonstrate that, in general, our Ciona optimized variants are detected and expressed at higher levels than commercially available fluorescent proteins. We show that three of these proteins, expressed simultaneously in different spatial domains within the same transgenic embryo are easily detectable using optimized fluorescent filter sets for epifluorescent microscopy. Coupled with recently developed quantitative imaging techniques, our GFP variants should provide useful reagents for monitoring the simultaneous expression of multiple genes in transgenic ascidian embryos. Developmental Dynamics 235:456,467, 2006. © 2005 Wiley-Liss, Inc. [source] Oligonucleotide-based microarray analysis of retinoic acid target genes in the protochordate, Ciona intestinalisDEVELOPMENTAL DYNAMICS, Issue 4 2005Tomoko Ishibashi Abstract Oligonucleotide-based microarray analyses were carried out to identify retinoic acid target genes in embryos of the ascidian Ciona intestinalis. Of 21,938 spots, 50 (corresponding to 43 genes) showed over twofold up-regulation in retinoic acid-treated tail bud embryos. In situ hybridization verified retinoic acid-induced up-regulation of 23 genes. Many of them were expressed in the anterior tail region, where a retinaldehyde dehydrogenase homolog is expressed. Homologs of vertebrate genes involved in neurogenesis and/or neuronal functions (e.g., COUP-TF, Ci-Hox1, and SCO-spondin) were expressed in the central nervous system of Ciona embryos, and activated by retinoic acid. Genes encoding transcription factors (e.g., Ci-lmx1.2, vitamin D receptor, and Hox proteins) and apoptosis-related proteins (e.g., transglutaminase and an apoptosis-inducing factor homolog) were also activated by retinoic acid. Simultaneous treatment of embryos with retinoic acid and puromycin revealed a few direct targets, including genes encoding Ci-Hox1, Ci-Cyp26, and an Rnf126-like ring finger protein. Developmental Dynamics 233:1571,1578, 2005. © 2005 Wiley-Liss, Inc. [source] Ci-GATAa, a GATA -class gene from the ascidian Ciona intestinalis: Isolation and developmental expressionDEVELOPMENTAL DYNAMICS, Issue 1 2003Palmira D'Ambrosio Abstract Members of the GATA family of zinc finger transcription factors have been shown to play important roles in controlling gene expression in a variety of cell types in many metazoan. Here, we describe the identification of Ci-GATAa, a member of this gene family, in the ascidian Ciona intestinalis. Whole-mount in situ hybridization showed that Ci-GATAa was expressed in a highly dynamic manner. The maternal transcript was evenly distributed in the embryo during early stages of development; however, the signal gradually decreased until it disappeared at the 64-cell stage. A zygotic transcript was detected at the 110-cell stage in the blastomeres precursors of three different tissues (brain vesicle, mesenchyme, and trunk lateral cells) and the signal was conserved in these territories up to the larval stage, indicating an important role for Ci-GATAa during ascidian differentiation. © 2002 Wiley-Liss, Inc. [source] Sequencing of real-world samples using a microfabricated hybrid device having unconstrained straight separation channelsELECTROPHORESIS, Issue 21 2003Shaorong Liu Abstract We describe a microfabricated hybrid device that consists of a microfabricated chip containing multiple twin-T injectors attached to an array of capillaries that serve as the separation channels. A new fabrication process was employed to create two differently sized round channels in a chip. Twin-T injectors were formed by the smaller round channels that match the bore of the separation capillaries and separation capillaries were incorporated to the injectors through the larger round channels that match the outer diameter of the capillaries. This allows for a minimum dead volume and provides a robust chip/capillary interface. This hybrid design takes full advantage, such as sample stacking and purification and uniform signal intensity profile, of the unique chip injection scheme for DNA sequencing while employing long straight capillaries for the separations. In essence, the separation channel length is optimized for both speed and resolution since it is unconstrained by chip size. To demonstrate the reliability and practicality of this hybrid device, we sequenced over 1000 real-world samples from Human Chromosome 5 and Ciona intestinalis, prepared at Joint Genome Institute. We achieved average Phred20 read of 675 bases in about 70 min with a success rate of 91%. For the similar type of samples on MegaBACE 1000, the average Phred20 read is about 550,600 bases in 120 min separation time with a success rate of about 80,90%. [source] Acetylcholinesterase from the invertebrate Ciona intestinalis is capable of assembling into asymmetric forms when co-expressed with vertebrate collagenic tail peptideFEBS JOURNAL, Issue 6 2008Adam Frederick To learn more about the evolution of the cholinesterases (ChEs), acetylcholinesterase (AChE) and butyrylcholinesterase in the vertebrates, we investigated the AChE activity of a deuterostome invertebrate, the urochordate Ciona intestinalis, by expressing in vitro a synthetic recombinant cDNA for the enzyme in COS-7 cells. Evidence from kinetics, pharmacology, molecular biology, and molecular modeling confirms that the enzyme is AChE. Sequence analysis and molecular modeling also indicate that the cDNA codes for the AChET subunit, which should be able to produce all three globular forms of AChE: monomers (G1), dimers (G2), and tetramers (G4), and assemble into asymmetric forms in association with the collagenic subunit collagen Q. Using velocity sedimentation on sucrose gradients, we found that all three of the globular forms are either expressed in cells or secreted into the medium. In cell extracts, amphiphilic monomers (G1a) and non-amphiphilic tetramers (G4na) are found. Amphiphilic dimers (G2a) and non-amphiphilic tetramers (G4na) are secreted into the medium. Co-expression of the catalytic subunit with Rattus norvegicus collagen Q produces the asymmetric A12 form of the enzyme. Collagenase digestion of the A12 AChE produces a lytic G4 form. Notably, only globular forms are present in vivo. This is the first demonstration that an invertebrate AChE is capable of assembling into asymmetric forms. We also performed a phylogenetic analysis of the sequence. We discuss the relevance of our results with respect to the evolution of the ChEs in general, in deuterostome invertebrates, and in chordates including vertebrates. [source] Images from the Woods Hole Summer of 2009 Embryology CourseGENESIS: THE JOURNAL OF GENETICS AND DEVELOPMENT, Issue 9 2009Article first published online: 24 SEP 200 Shown are images of Drosophila melanogaster (Fruit fly), Xenopus laevis (African clawed frog), Schmidtea mediterranea (Planaria), Hydroides (Serpulid worm), Schistocerca americana (American bird grasshopper), Euprymna scolopes (Hawaiian bobtail squid), Ciona intestinalis (Vase tunicate), Phalangium opilio (Daddy longlegs), Artemia franciscana (Brine shrimp), Mustelus canis (Dogfish), Danio rerio (Zebrafish), Gallus gallus domesticus (Chicken), Mnemiopsis leidyi (Warty comb jelly), Oscarella carmela (Desmosponge), Chaetopterus variopedatus (Parchment worm), and the Marbled crayfish that were generated and taken by members of the Woods Hole Embryology Course in the summer of 2009. Photo credits: Neel Aluru, Otger Campas, Carlos Carmona-Fontaine, Sheng-hong Chen, Katrien De Mulder, April Dinwiddie, Adele M. Doyle, Antje Fischer, Claudiu Giurumescu, Lauretta Grasso, Alysha Heimberg, Francie Hyndman, Erin Kaltenbrun, Dov Lerman-Sinkoff, Dede Lyons, Chema Martin-Durán, Lara Marxreiter, Jeremy Mosher, Malea Murphy, Lee Niswander, Vincent Pasque, Nipam H. Patel, Alberto Roselló, Prashant Sharma, Ashley Siegel, Ajay Thomas, Frank Tulenko, Alex Vasilyev, and Naveen Wijesena. For more information on the Embryology Course, please visit http://www.mblembryology.org/. [source] High-throughput enhancer trap by remobilization of transposon Minos in Ciona intestinalisGENESIS: THE JOURNAL OF GENETICS AND DEVELOPMENT, Issue 5 2007Satoko Awazu Abstract The enhancer trap approach utilizing transposons yields us information about gene functions and gene expression patterns. In the ascidian Ciona intestinalis, transposon-based transgenesis and insertional mutagenesis were achieved with a Tc1/mariner transposon Minos. We report development of a novel technique for enhancer trap in C. intestinalis. This technique uses remobilization of Minos in the Ciona genome. A Minos vector for enhancer trap was constructed and a tandem array insertion of the vector was introduced into the Ciona genome to create a mutator line. Minos was remobilized in Ciona chromosomes to create new insertions by providing transposases. These transposase-introduced animals were crossed with wild-type animals. Nearly 80% of F1 families showed novel GFP expression patterns. This high-throughput enhancer trap screen will be useful to create new marker transgenic lines showing reporter gene expression in specific tissues and to identify novel patterns of gene expression. genesis 45:307,317, 2007. © 2007 Wiley-Liss, Inc. [source] Characterization of novel GPCR gene coding locus in amphioxus genome: Gene structure, expression, and phylogenetic analysis with implications for its involvement in chemoreceptionGENESIS: THE JOURNAL OF GENETICS AND DEVELOPMENT, Issue 2 2005Gouki Satoh Abstract Chemosensation is the primary sensory modality in almost all metazoans. The vertebrate olfactory receptor genes exist as tandem clusters in the genome, so that identifying their evolutionary origin would be useful for understanding the expansion of the sensory world in relation to a large-scale genomic duplication event in a lineage leading to the vertebrates. In this study, I characterized a novel GPCR (G-protein-coupled receptor) gene-coding locus from the amphioxus genome. The genomic DNA contains an intronless ORF whose deduced amino acid sequence encodes a seven-transmembrane protein with some amino acid residues characteristic of vertebrate olfactory receptors (ORs). Surveying counterparts in the Ciona intestinalis (Asidiacea, Urochordata) genome by querying BLAST programs against the Ciona genomic DNA sequence database resulted in the identification of a remotely related gene. In situ hybridization analysis labeled primary sensory neurons in the rostral epithelium of amphioxus adults. Based on these findings, together with comparison of the developmental gene expression between amphioxus and vertebrates, I postulate that chemoreceptive primary sensory neurons in the rostrum are an ancient cell population traceable at least as far back in phylogeny as the common ancestor of amphioxus and vertebrates. genesis 41:47,57, 2005. © 2005 Wiley-Liss, Inc. [source] Protease,proteoglycan complexes of mouse and human mast cells and importance of their ,-tryptase,heparin complexes in inflammation and innate immunityIMMUNOLOGICAL REVIEWS, Issue 1 2007Richard L. Stevens Summary:, Approximately 50% of the weight of a mature mast cell (MC) consists of varied neutral proteases stored in the cell's secretory granules ionically bound to serglycin proteoglycans that contain heparin and/or chondroitin sulfate E/diB chains. Mouse MCs express the exopeptidase carboxypeptidase A3 and at least 15 serine proteases [designated as mouse MC protease (mMCP) 1,11, transmembrane tryptase/tryptase ,/protease serine member S (Prss) 31, cathepsin G, granzyme B, and neuropsin/Prss19]. mMCP-6, mMCP-7, mMCP-11/Prss34, and Prss31 are the four members of the chromosome 17A3.3 family of tryptases that are preferentially expressed in MCs. One of the challenges ahead is to understand why MCs express so many different protease,proteoglycan macromolecular complexes. MC-like cells that contain tryptase,heparin complexes in their secretory granules have been identified in the Ciona intestinalis and Styela plicata urochordates that appeared approximately 500 million years ago. Because sea squirts lack B cells and T cells, it is likely that MCs and their tryptase,proteoglycan granule mediators initially appeared in lower organisms as part of their innate immune system. The conservation of MCs throughout evolution suggests that some of these protease,proteoglycan complexes are essential to our survival. In support of this conclusion, no human has been identified that lacks MCs. Moreover, transgenic mice lacking the ,-tryptase mMCP-6 are unable to combat a Klebsiella pneumoniae infection effectively. Here we summarize the nature and function of some of the tryptase,serglycin proteoglycan complexes found in mouse and human MCs. [source] Identification of testis-specific ubiquitin-conjugating enzyme in the ascidian Ciona intestinalisMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 7 2010Naoto Yokota The ubiquitin,proteasome system is known to play a key role in fertilization in ascidians, sea urchins, and mammals. To obtain insights into the ubiquitin-conjugating enzymes (Ube2) involved in reproductive systems, we systematically explored Ube2 enzymes expressed in the testis of the ascidian Ciona intestinalis. Here, we report cDNA cloning and characterization of a novel type of Ube2r (Ci0100152677) that is capable of making a thiolester bond with ubiquitin. Northern analysis, whole-mount in situ hybridization and immunocytochemistry indicate that this enzyme is exclusively expressed in the testis, mainly in the germ cells during the late stage of spermatogenesis, and is localized in the sperm head and tail, suggesting possible participation in fertilization or spermatogenesis/spermiogenesis. Mol. Reprod. Dev. 77: 640,647, 2010. © 2010 Wiley-Liss, Inc. [source] Ion current activity and molecules modulating maturation and growth stages of ascidian (Ciona intestinalis) oocytesMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 11 2009Francesco Silvestre Electrophysiological techniques were used to study ion currents in the ascidian Ciona intestinalis oocyte plasma membranes during different stages of growth and meiosis. Three stages (A, B, C) of immature oocytes were discriminated in the ovary, with the germinal vesicle (GV) showing specific different features of growth and maturation. Stage A (pre-vitellogenic) oocytes exhibited the highest L-type Ca2+current activity, and were incompetent for meiosis resumption. Stage B (vitellogenic) oocytes showed Na+ currents that remained high during the maturation, up to the post-vitellogenic stage C oocytes. The latter had acquired meiotic competence, undergoing spontaneous maturation and interacting with the spermatozoon. However, fertilized oocytes did not produce normal larvae, suggesting that cytoplasmic maturation plays a specific role in embryo development. Spontaneous maturation was inhibited at low pH whereas trypsin was able to trigger germinal vesicle breakdown (GVBD) regardless of pH; in addition spontaneous maturation was not affected by removal of follicle cells or by inhibiting junctional communication between oocyte and follicle cells. Taken together these results imply: (i) Ca2+ and Na+ currents are involved in meiotic progression, growth, and acquisition of meiotic competence; (ii) trypsin-like molecules may have a role as candidates for providing the physiological stimulus to resume meiosis. Finally, we provide evidence that follicle cells in Ciona are not involved in triggering GVBD as it occurs in other ascidians. Mol. Reprod. Dev. 76: 1084,1093, 2009. © 2009 Wiley-Liss, Inc. [source] Bioactive aldehydes from diatoms block the fertilization current in ascidian oocytesMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 1 2003Elisabetta Tosti Abstract The effects of bioactive aldehydes from diatoms, unicellular algae at the base of the marine food web, were studied on fertilization and early development processes of the ascidian Ciona intestinalis. Using whole-cell voltage clamp techniques, we show that 2- trans -4- trans -decadienal (DD) and 2- trans -4- cis -7- cis -decatrienal (DT) inhibited the fertilization current which is generated in oocytes upon interaction with the spermatozoon. This inhibition was dose-dependent and was accompanied by inhibition of the voltage-gated calcium current activity of the plasma membrane. DD and DT did not inhibit the subsequent contraction of the cortex. Moreover, DD specifically acted as a fertilization channel inhibitor since it did not affect the steady state conductance of the plasma membrane or gap junctional (GJ) communication within blastomeres of the embryo. On the other hand, DD did affect actin reorganization even though the mechanism of action on actin filaments differed from that of other actin blockers. Possibly this effect on actin reorganization was responsible for the subsequent teratogenic action on larval development. The effect of DD was reversible if oocytes were washed soon after fertilization indicating that DD may specifically target certain fertilization mechanisms. Thus, diatom reactive aldehydes such as DD may have a dual effect on reproductive processes, influencing primary fertilization events such as gating of fertilization channels and secondary processes such as actin reorganization which is responsible for the segregation of cell lineages. These findings add to a growing body of evidence on the antiproliferative effects of diatom-derived aldehydes. Our results also report, for the first time, on the action of a fertilization channel blocker in marine invertebrates. Mol. Reprod. Dev. 66: 72,80, 2003. © 2003 Wiley-Liss, Inc. [source] Origin of the Vertebrate Visual Cycle: III.PHOTOCHEMISTRY & PHOTOBIOLOGY, Issue 6 2006-Monooxygenase Homologues in Ciona intestinalis, -carotene 1, Distinct Distribution of RPE6 We previously identified three genes that encode putative visual cycle proteins that are homologues of retinal G-protein coupled receptor (Ci-opsin3), cellular retinaldehyde-binding protein (Ci-CRALBP) and ,-carotene 15,15,-monooxygenase (Ci-BCO) in the ascidian Ciona intestinalis. Ci-opsin3 and Ci-CRALBP are localized in both ocellus photoreceptor cells and surrounding non-photoreceptor cells in the brain vesicle of the larva. In the present study, we investigated the possible role and evolutionary origin of the BCO/RPE65 family in the visual cycle by analyzing Ci-BCO localization by immunohistochemistry and by identifying a novel gene that encodes a homologue of retinal pigment epithelium,specific 65 kDa protein (Ci-RPE65) in C. intestinalis. In situ hybridization and expressed sequence tag (EST) profiles consistently suggest that Ci-RPE65 is not significantly expressed in the ocellus and brain vesicle of the larva. Ci-RPE65 is expressed in the neural complex, a photoreceptor organ of the adult ascidian, at a level comparable to that of Ci-opsin3 and Ci-CRALBP. Ci-RPE65 is also expressed in various adult tissues, including the gill, body wall and intestine, suggesting that Ci-RPE65 plays a role in addition to that in the visual cycle. In contrast, Ci-BCO is predominantly localized in ocellus photoreceptor cells of the larva. The larval visual cycle seems to use Ci-opsin3 as a photoisomerase. Our results also suggest that the RPE65-dependent visual cycle is used in the adult photoreceptors of a primitive chordate. [source] | |||||||||||||