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At least 19 recordsLinked to original sources
PubMed2000

Mechanism of the block to hybridization and selfing between the sympatric ascidians Ciona intestinalis and Ciona savignyi.

The solitary ascidians Ciona intestinalis and Ciona savignyi co-occur in southern California harbors, but no hybrids have been recognized in nature. Numerous differences in their egg morphology were detected. Homologous (normal outcross) fertilization yielded 96-99% cleavage, where autologous (self) fertilization showed 3% and heterologous (hybrid) fertilization showed 0-1%. Acid treatment (pH 3.2) removed the block to selfing (P < 0.0001) but not hybridization for both species. Heterologous sperm bind to the vitelline coat (VC), but fail to penetrate. Enzymatic removal of the VC resulted in 91-97% cleavage with autologous and heterologous sperm (P < 0.0001). The vitelline coats of the two species differ in lectin binding to surface glycosides. Fertilization in both species is significantly inhibited by the lectins, fucose binding protein (P < 0.0001) and concanavalin A (P < 0.0001), and wheat germ agglutinin inhibits fertilization in C. intestinalis (P < 0.0001) but is without effect on C. savignyi fertilization. Self and hybrid blocks employ different mechanisms including glycoside composition and acid sensitivity.

Animals↗
PubMed2006

Embryonic expression profiles and conserved localization mechanisms of pem/postplasmic mRNAs of two species of ascidian, Ciona intestinalis and Ciona savignyi.

In many animals, the first cue for development is transcripts and/or proteins that are provided maternally and are localized at specific regions of fertilized eggs and early embryos. The ascidian is known to exhibit a mosaic mode of development, which is largely dependent on localized maternal factors. In early Ciona intestinalis embryos, the posterior-most localization appears to be the major specialized pattern of maternal transcripts. The present study examined the temporal and spatial expression pattern of 40 genes known as pem/postplasmic genes, for which maternal mRNAs are localized at the posterior-most region during early Ciona embryogenesis. Ten of these genes showed redistribution to B8.12-line cells, which are known to give rise to germ cells in ascidians. In addition 23 orthologues were newly identified in a related ascidian species, Ciona savignyi, and 16 of them showed the mRNA localization pattern at the posterior-most region. Furthermore, the localized pattern of exogenous mRNA, which comprised the 3' UTR of C. intestinalis pem/postplasmic genes conjugated with the LacZ ORF, showed the localization at the posterior-most region in C. savignyi embryos. Likewise, the 3' UTR of C. savignyi pem/postplasmic genes conjugated with the LacZ ORF showed localization at the posterior most region in C. intestinalis embryos, suggesting that localization mechanisms are conserved between the two species. The present study therefore provides basic information for future functional analyses of these pem/postplasmic genes and for exploring the mechanisms of localization of mRNAs.

Animals↗
PubMed2006

Three insulin-relaxin-like genes in Ciona intestinalis.

The Ciona intestinalis genome harbors three insulin-like genes: INS-L1, -L2 and -L3. Conserved synteny between the Ciona-human genomes predicts that Ciona INS-Ls are orthologous to the vertebrate insulin-relaxin family, but this relation cannot be inferred from molecular phylogeny. A conserved protein core with six cysteines; typical arrangement of B-, C- and A-protein domains; pro-protein maturation mode; and putative insulin receptor-binding sites were identified in Ciona INS-L proteins. ESTs used to assemble exonic sequences of INS-Ls combined with qRT-PCR analysis provided evidence that the predicted genes are expressed in the developing and adult Ciona. Our results support that Ciona INS-L1 is orthologous to the vertebrate insulin-like/relaxin genes, INS-L2 to insulin genes and INS-L3 to IGF genes. Our analysis also implies that the insulin-like/relaxin ancestor switched receptor type from tyrosine kinase- to GPCR-type, whereas insulin-IGF subfamily retained the tyrosine kinase-type of receptor. We propose that this receptor-switch occurred after the time when urochordates branched from the common chordate lineage, but before the two genome-duplications at the root of the vertebrates.

Amino Acid Sequence↗
PubMed1990

Chymotrypsin-like enzymes are involved in sperm penetration through the vitelline coat of Ciona intestinalis egg.

In Ciona intestinalis, sperm penetration through the egg vitelline coat is an essential event of fertilization. We investigated whether trypsin- and chymotrypsin-like enzymes are involved in this event. Inhibitors and peptide substrates for chymotrypsin-like enzymes blocked the overall process of fertilization in a concentration-dependent manner. The inhibitory activity was specifically exerted on the step of sperm penetration. Chymotrypsin-like protease activity was identified in spermatozoa with the fluorogenic synthetic substrate Suc-Ala-Ala-Phe-AMC, which was the most effective substrate in blocking sperm penetration. These data indicate that a chymotrypsin-like protease activity is a sperm lysin of Ciona intestinalis.

Amino Acid Sequence↗
PubMed2005

Cataloging transcription factor and major signaling molecule genes for functional genomic studies in Ciona intestinalis.

The ascidian Ciona intestinalis provides an excellent experimental system for functional genomic studies because (1) its genome has been sequenced, (2) the transcription factor genes and genes for major signal transduction molecules have been extensively screened and annotated on a genome-wide scale using the molecular phylogenetical method, and (3) their embryonic expression profiles have been almost completely determined. However, the entire genetic structure, including the 5' and 3' untranslated regions and the protein-coding regions, of most gene models used in these prior studies is not always supported by cDNA evidence, and thus, these gene models are potentially imprecise. To facilitate functional genomic studies based on precise gene structures, our present study determined 406 cDNA sequences for 357 transcription factor genes and 112 cDNA sequences for 107 signal transduction molecule genes, greatly improving the previous gene models and revealing transcript variants for 44 genes. Considering these data alongside those of previously characterized genes deposited in the DNA Data Bank of Japan/European Molecular Biology Laboratory/GENBANK databases, 95.6% of the catalogued transcription factor genes (373/390) and 98.3% of the catalogued signal transduction molecule genes (117/119) have now been verified by cDNA sequences. Thus, the present study greatly improves the resources available for functional genomic studies in C. intestinalis.

Animals↗
PubMed2005

Live imaging and morphometric analysis of embryonic development in the ascidian Ciona intestinalis.

The ascidian Ciona intestinalis is one of the model organisms of choice for comparative investigations of chordate development and for unraveling the molecular mechanisms underlying morphogenesis and cell fate specification. Taking advantage of the availability of various genetically encoded fluorescent proteins and of defined cis-regulatory elements, we combined transient transgenesis with laser scanning confocal imaging to acquire and quantitate 3D time-lapse data from living Ciona embryos. We used Ciona tissue-specific enhancers to drive expression of spectrally distinct fluorescent protein reporters to label and simultaneously visualize axially and paraxially positioned mesodermal derivatives, as well as neural precursors in individual embryos. We observed morphogenetic movements, without perturbing development, from the early gastrula throughout the larval stage, including gastrulation, neurulation, convergent extension of the presumptive notochord, and tail elongation. These multidimensional data allowed us to establish a reference system of metrics to quantify key developmental events including blastopore closure and muscle extension. The approach we describe can be used to document morphogenetic cell and tissue rearrangements in living embryos and paves the way for a live digitized anatomical atlas of Ciona.

Animals↗
PubMed1992

Purification and characterization of a vitelline coat lysin from Ciona intestinalis spermatozoa.

In Ciona intestinalis a chymotrypsin-like activity is involved in sperm penetration of the egg vitelline coat. A chymotrypsin-like enzyme has been purified from spermatozoa by a protocol including ion exchange chromatography, gel filtration, and native polyacrylamide gel electrophoresis. The purified enzyme resulted homogeneous when analyzed by SDS-PAGE. The molecular weight of the chymotrypsin-like enzyme was estimated to be 35 kDa by gel filtration and 24 KDa by SDS-PAGE in nonreducing conditions. The pH optimum of the enzyme is 8.4 and its activity is enhanced by Ca2+. It shows the highest activity towards the synthetic substrate Suc-Ala-Ala-Pro-Phe-AMC. Furthermore, by electron microscopy, the purified enzyme affects the structure of egg vitelline coat, and thus it fulfills one of the criteria of a lysin.

Amino Acid Sequence↗
PubMed1997

Specific cellular localization of tyrosinase mRNA during Ciona intestinalis larval development.

A Ciona intestinalis cDNA clone that encodes a protein highly homologous to other tyrosinases was isolated. Northern blot analysis showed that expression of Ciona tyrosinase starts at the early neurula stage and continues throughout the tail-bud and tadpole larval stages. The earliest tyrosinase expression was detected, by in situ hybridization, at the neural plate stage, in pigment precursor cells located along the two neural folds, in the animal region of the embryo. In the course of embryonic development the strong hybridization signal was always localized, within the rostral part of the developing brain, in the pigment precursor cells and was later detected in the otolith and ocellus. These results are discussed in relation to tyrosinase as an early marker of neural induction.

Amino Acid Sequence↗
PubMed1999

Membrane hyperpolarization by sperm-activating and -attracting factor increases cAMP level and activates sperm motility in the ascidian Ciona intestinalis.

In the ascidian Ciona intestinalis (and C. savignyi), sperm-activating and -attracting factor (SAAF) is released from the egg at fertilization and stimulates both Ca(2+) influx and a transient increase in cAMP level of the sperm, leading to the activation of sperm motility (M. Yoshida et al., 1994, Dev. Growth Differ. 36, 589-595). In this paper we show in C. intestinalis that valinomycin, a potassium-selective ionophore, as well as SAAF, activated sperm motility, and this activation was suppressed by extracellular high K(+). Membrane potential measurements showed that both SAAF and valinomycin increase K(+) permeability of sperm and induce membrane hyperpolarization, the amplitude of which depends on the external K(+) concentration. The membrane potential and intracellular K(+) concentration of Ciona sperm without SAAF were estimated to be about -50 mV and 560 +/- 40 mM, respectively. After treatment with SAAF or valinomycin the membrane potential became almost equal to the equilibrium potential of K(+) (-100 mV), and the cAMP level increased in artificial seawater. A potent voltage-dependent K(+) channel blocker, MCD peptide, at the concentration of 10 microM blocked SAAF-induced hyperpolarization of the cells, increase in cAMP, and sperm motility. These results suggest that membrane hyperpolarization produced by the opening of K(+) channels elevates cAMP synthesis and leads to the activation of sperm motility in Ciona.

Animals↗
PubMed2005

Genomic approaches reveal unexpected genetic divergence within Ciona intestinalis.

The invertebrate chordate Ciona intestinalis is a widely used model organism in biological research. Individuals from waters ranging from arctic to temperate are morphologically almost indistinguishable. However, we found significant differences in whole genomic DNA sequence between northern European and Pacific C. intestinalis. Intronic and transposon sequences often appear unrelated between these geographic origins and amino acid substitutions in protein coding sequences indicate a divergence time in excess of 20 MYA. This finding suggests the existence of two cryptic species within the present C. intestinalis species. We found five marker loci which distinguish the two genetic forms by PCR. This analysis revealed that specimens from Naples, Italy, have the Pacific-type genome, perhaps due to human-mediated marine transport of species. Despite major genomic divergence, the two forms could be hybridized in the laboratory.

Animals↗
PubMed2003

Construction of a cDNA microarray derived from the ascidian Ciona intestinalis.

A cDNA microarray was constructed from a basal chordate, the ascidian Ciona intestinalis. The draft genome of Ciona has been read and inferred to contain approximately 16,000 protein-coding genes, and cDNAs for transcripts of 13,464 genes have been characterized and compiled as the "Ciona intestinalis Gene Collection Release I". In the present study, we constructed a cDNA microarray of these 13,464 Ciona genes. A preliminary experiment with Cy3- and Cy5-labeled probes showed extensive differential gene expression between fertilized eggs and larvae. In addition, there was a good correlation between results obtained by the present microarray analysis and those from previous EST analyses. This first microarray of a large collection of Ciona intestinalis cDNA clones should facilitate the analysis of global gene expression and gene networks during the embryogenesis of basal chordates.

Animals↗
PubMed2002

A cDNA resource from the basal chordate Ciona intestinalis.

The genome of the basal choradate Ciona intestinalis contains a basic set of genes with less redundancy compared to the vertebrate genome. Extensive EST analyses, cDNA sequencing, and clustering yielded "Ciona intestinalis Gene Collection Release 1," which contains cDNA clones for 13,464 genes, covering nearly 85% of the Ciona mRNA species. This release is ready for use in cDNA cloning, micro/macroarray analysis, and other comprehensive genome-wide analyses for further molecular studies of basal chordates.

Animals↗
PubMed1990

Temporal and spatial correlation of fertilization current, calcium waves and cytoplasmic contraction in eggs of Ciona intestinalis.

Eggs of the ascidian Ciona intestinalis were loaded with the calcium indicator fura-2 via whole-cell clamp electrodes and changes in cytoplasmic calcium and cell currents were monitored during fertilization either in separate eggs or simultaneously in the same egg. The first indication of egg activation was the fertilization current; which reached peak values around 1 nA after 30 s. A wave of elevated calcium was detectable between 5 s and 30 s (mean = 21 s) after the start of the fertilization current. This wave spread across the egg increasing cytoplasmic calcium levels to at least 10 microM. When the fertilization current and calcium wave were complete and cytoplasmic calcium levels were decreasing to prefertilization levels, a cortical contraction wave spread across the egg surface. In eggs showing normal fertilization current, the calcium wave and the contraction wave were in the same direction. A region of elevated calcium persisted at the animal pole. Changing cytoplasmic calcium levels locally by local application of ionophore A23187 caused a contraction wave originating at the site of ionophore application. Increasing cytoplasmic calcium uniformly by facilitating calcium entry through voltage-regulated channels did not result in a contraction wave.

Animals↗
PubMed1998

The hsp70 protein is involved in the acquisition of gamete self-sterility in the ascidian Ciona intestinalis.

In the hermaphrodite ascidian Ciona intestinalis, gamete self-incompatibility is a mechanism that prevents self-fertilization and is based on the ability of the oocyte vitelline coat to distinguish and accept only heterologous spermatozoa. The onset of self-sterility occurs during oogenesis and involves or is controlled by the follicle cells. Gamete self-nonself discrimination, a process that can be likened to an immune recognition event, represents a useful model with which to study the evolution of self-nonself recognition. Hsp70 genes, which belong to the major histocompatibility complex (MHC) class III, are supposedly ancestors of the MHC class I and II genes, and chaperonins are known to be involved in antigen processing and presentation. We have isolated and characterized an hsp70 gene (Cihsp70) that is constitutively expressed during oogenesis in the follicle cells of previtellogenic and vitellogenic oocytes. Using a polyclonal antibody against Cihsp70 protein, we demonstrate that the expression of Cihsp70 is required for the switch from self-fertility to self-sterility. The functional involvement of Cihsp70 in gamete self-nonself recognition provides evidence for an ancestral MHC-like system in protochordates.

Amino Acid Sequence↗
PubMed2006

FGF signaling delineates the cardiac progenitor field in the simple chordate, Ciona intestinalis.

Comprehensive gene networks in Ciona intestinalis embryos provide a foundation for characterizing complex developmental processes, such as the initial phases of chordate heart development. The basic helix-loop-helix regulatory gene Ci-Mesp is required for activation of cardiac transcription factors. Evidence is presented that Ci-Ets1/2, a transcriptional effector of receptor tyrosine kinase (RTK) signaling, acts downstream from Mesp to establish the heart field. Asymmetric activation of Ets1/2, possibly through localized expression of FGF9, drives heart specification within this field. During gastrulation, Ets1/2 is expressed in a group of four cells descended from two Mesp-expressing founder cells (the B7.5 cells). After gastrulation, these cells divide asymmetrically; the smaller rostral daughters exhibit RTK activation (phosphorylation of ERK) and form the heart lineage while the larger caudal daughters form the anterior tail muscle lineage. Inhibition of RTK signaling prevents heart specification. Targeted inhibition of Ets1/2 activity or FGF receptor function also blocks heart specification. Conversely, application of FGF or targeted expression of constitutively active Ets1/2 (EtsVp16) cause both rostral and caudal B7.5 lineages to form heart cells. This expansion produces an unexpected phenotype: transformation of a single-compartment heart into a functional multicompartment organ. We discuss these results with regard to the development and evolution of the multichambered vertebrate heart.

Animals↗