Ciona: A Model for Developmental Genomics

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Ciona: A Model for Developmental Genomics

Noriyuki Satoh, Okinawa Institute of Science and Technology, Okinawa, Japan

Published online: March 2013

DOI: 10.1002/9780470015902.a0021411

Full Article on Wiley Online Library

Abstract
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Abstract

The ascidian Ciona intestinalis is an excellent model system for studying developmental biology due to the simplicity of its embryogenesis and genomics. The Ciona tadpole larva is composed of only 2600 cells and possesses the basic body plan of chordates. The 117‐Mbp‐long euchromatic genome encodes approximately 320 core transcription factors and 125 major signal transduction molecules, the expression profiles of which have been described at the single‐cell level in Ciona embryos. Not only the function of developmentally relevant genes can be explored by means of suppressed and/or ectopic expression, but also a convenient electroporation method of reporter gene constructs has been developed to identify cis‐regulatory modules. High throughput and genome‐wide approaches can be used to uncover gene regulatory networks involved in cell specification and differentiation. Transposon‐mediated transgenesis also provides a new strategy for elucidating the mechanisms of gene expression and function. These and other advantages of the Ciona system recommend future developmental biology studies with this model.

Key Concepts:

The Ciona tadpole larva represents the basic and most simplified chordate body plan, and its embryogenesis is well characterised.
The Ciona intestinalis genome has been decoded. The 117‐Mbp euchromatic genome, which is packaged into 14 pairs of chromosomes, contains 15 254 genes including ∼320 core TF genes and 125 major signalling molecule genes.
Most of these developmentally relevant genes have been chromosomally mapped.
The spatial and temporal expression profiles of these developmentally relevant genes have been elucidated.
Gene function can be easily studied by suppressed and/or ectopic expression.
A convenient electroporation method allows quantitative and qualitative examination of cis‐regulatory modules responsible for gene expression.
The regulatory networks of TF and signalling molecule genes that establish the blueprint of the chordate body plan have been uncovered.
The short generation time allows forward genetics of Ciona, a first for a marine invertebrate.
Transgenic lines have been established using Minos transposons to reveal gene function and expression.

Keywords: ascidian; Ciona intestinalis; tadpole larvae; basic chordate body plan; decoded genome; transcriptomics; forward genetics; transcription factors; cell–cell signalling molecules; cis‐regulatory modules; gene regulatory network

	Figure 1. The ascidian urochordate C. intestinalis. (a) Adults with incurrent and outcurrent syphons. The white duct is the sperm duct and the parallel orange duct is the egg duct. (b–g) Embryogenesis. Embryos were dechorionated to show their outer morphology clearly: (b) Fertilised egg, (c) 16‐cell embryo, (d) gastrula (approximately 150 cells), (e, f) tailbud embryos and (g) tadpole larva. (h) A juvenile, a few days after metamorphosis, showing the internal structures: ds, digestive system; en, endostyle: ht, heart; os, neuronal complex; and pg, pharyngeal gill. (i) Fate map of the 110‐cell embryo, animal hemisphere (left) and vegetal hemisphere (right). Blastomeres are named according to Conklin's nomenclature and coloured according to developmental fate restriction. Green, epidermis; dark blue, brain; light blue, nerve cord; magenta, notochord; light blue, trunk lateral cells; dark green, mesenchyme; yellow, endoderm; and red, muscle. (j) Oblique lateral confocal section through a mid‐tailbud stage C. savignyi embryo. The 40 notochord cells (green) were marked with a stable Brachyury:GFP transgene. Cell peripheries were labelled with phalloidin and manually pseudocoloured to show the endoderm (blue), muscle (red), neural tube (yellow) and epidermis (magenta). CNS, central nervous system; End, endoderm; Epi, epidermis; NC, nerve cord; Mch, mesenchyme; Mu, muscle; Not, notochord. Reproduced with permission from ‘Development’ 1 January 2008, 135 (1) (cover image) Image: William Smith.
	Figure 2. Chromosomal map of 373 core TF genes and 111 major cell signalling molecule genes in C. intestinalis. Families of TF are shown by different coloured discs, whereas families of cell signalling molecules are shown by different coloured arrowheads (bottom right). Centromeric regions are shown by dark blue dashed lines. Red dashed lines indicate three rDNA cluster regions and green dashed lines indicate a histone cluster region. Blue dashed lines indicate unmapped regions. The left and right vertical lines of each chromosome indicate the 5′–3′ and 3′–5′ alignment, respectively. The telomeric regions on the short arms of chromosomes 12, 13 and 14 are ordered arbitrarily.
	Figure 3. A method to examine regulatory modules responsible for specific gene expression by electroporation of reporter gene constructs. (a) Fertilised or unfertilised eggs (brown circles) within an electroporation chamber filled with seawater containing reporter gene constructs (black bars). (b) Culture of manipulated embryos in agar‐coated plastic dishes. (c) Detection of reporter (lacZ or GFP) expression in larvae. In this case, GFP expression driven by Brachyury promoter is evident. This process (from a to c) only takes ∼20 h. Reproduced with permission from: Cover image from Science vol. 298, no. 5601, 13 December 2002. Image: Mei Wang. © American Association for the Advancement of Science.
	Figure 4. Regulatory network of zygotically expressed TF and STM genes in the early Ciona embryo. (a) A 64‐cell stage embryo viewed from the animal (upper right) or vegetal (lower right) pole, with nomenclature of blastomeres, and the line of embryonic cells (left) indicated. (b) Expression patterns of 36 regulatory genes zygotically expressed at the 64‐cell stage. (c) The regulatory network of 76 genes expressed in early embryos, up to the early gastrula stage. The genes are ordered alphabetically from the top‐left corner to the bottom‐right corner. The chromosomal localisation of the genes is also shown in light boxes, with short (left) and long (right) arms. Lines show the gene regulatory networks, dark blue lines and arrows indicate the same chromosomal interactions, orange lines show interchromosomal interactions and green arrows indicate autoregulatory interactions.
	Figure 5. Transgenic Ciona lines. (a) Production of transgenic lines with specific markers. Unfertilised eggs are first microinjected with transposon vector including GFP and transposase mRNA. They are then inseminated and allowed to develop into mature adults. Founder sperm is used to fertilise eggs of wild‐type animals. These eggs are then cultured to larval, juvenile and adult stages, from which specimens with GFP expression are selected. (b–f) Examples of transgenic lines with specific markers: (b) A transgenic line in which GFP is specifically expressed in the epidermis, (c) notochord, (d) muscle or (e) central nervous system of larva. (f) A line in which juvenile expresses GFP in the nervous system and red fluorescent protein in the endostyle. Courtesy of Dr. Sasakura.

Article Title:	Ciona: A Model for Developmental Genomics
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