Abstract
Ascidian embryos have been employed as model systems for studies of developmental biology for well over a century, owing to their desirable blend of experimental advantages, which include their rapid development, traceable cell lineage, and evolutionarily conserved morphogenetic movements. Two decades ago, the development of a streamlined electroporation method drastically reduced the time and cost of transgenic experiments, and, along with the elucidation of the complete genomic sequences of several ascidian species, propelled these simple chordates to the forefront of the model organisms available for studies of regulation of gene expression. Numerous ascidian sequences with tissue-specific enhancer activity were isolated and rapidly characterized through systematic in vivo experiments that would require several weeks in most other model systems. These cis-regulatory sequences include a large collection of notochord enhancers, which have been used to visualize notochord development in vivo, to generate mutant phenotypes, and to knock down genes of interest. Moreover, their detailed characterization has allowed the reconstruction of different branches of the notochord gene regulatory network. This chapter describes how the use of transgenic techniques has rendered the ascidian Ciona a competitive model organism for studies of notochord development, evolution, and gene regulation.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Abbreviations
- bp:
-
Base pair(s)
- cDNA:
-
Complementary DNA
- ChIP:
-
Chromatin immunoprecipitation
- CRM:
-
cis-regulatory module
- DAPI:
-
4′,6-diamidino-2-phenylindole
- FACS:
-
fluorescence-activated cell sorting
- GFP:
-
Green fluorescent protein
- GRN:
-
Gene regulatory network
- NOCE:
-
Notochord enhancer
- OBS:
-
Orphan binding site
- P3H1:
-
prolyl 3-hydroxylase1
- shRNA:
-
Short hairpin RNA
References
Adams RR, Tavares AA, Salzberg A, Bellen HJ, Glover DM (1998) Pavarotti encodes a kinesin-like protein required to organize the central spindle and contractile ring for cytokinesis. Genes Dev 12(10):1483–1494
Alten L, Schuster-Gossler K, Eichenlaub MP, Wittbrodt B, Wittbrodt J, Gossler A (2012) A novel mammal-specific three partite enhancer element regulates node and notochord-specific Noto expression. PLoS One 7(10):e47785
Ang SL, Rossant J (1994) HNF-3 beta is essential for node and notochord formation in mouse development. Cell 78:561–574
Anno C, Satou A, Fujiwara S (2006) Transcriptional regulation of ZicL in the Ciona intestinalis embryo. Dev Genes Evol 216(10):597–605
Bagheri-Fam S, Barrionuevo F, Dohrmann U, Günther T, Schüle R, Kemler R, Mallo M, Kanzler B, Scherer G (2006) Long-range upstream and downstream enhancers control distinct subsets of the complex spatiotemporal Sox9 expression pattern. Dev Biol 291(2):382–397
Capellini TD, Dunn MP, Passamaneck YJ, Selleri L, Di Gregorio A (2008) Conservation of notochord gene expression across chordates: insights from the Leprecan gene family. Genesis 46(11):683–696
Casey ES, O'Reilly MA, Conlon FL, Smith JC (1998) The T-box transcription factor Brachyury regulates expression of eFGF through binding to a non-palindromic response element. Development 125(19):3887–3894
Chiba S, Jiang D, Satoh N, Smith WC (2009) Brachyury null mutant-induced defects in juvenile ascidian endodermal organs. Development 136(1):35–39
Christiaen L, Davidson B, Kawashima T, Powell W, Nolla H, Vranizan K, Levine M (2008) The transcription/migration interface in heart precursors of Ciona intestinalis. Science 320(5881):1349–1352
Cleaver O, Krieg PA (2001) Notochord patterning of the endoderm. Dev Biol 234(1):1–12
Cloney RA (1964) Development of the ascidian notochord. Acta Embryol Morphol Exp 7:111–130
Conklin EG (1905) The organization and cell-lineage of the ascidian egg. J Acad Nat Sci 13:1–119
Corbo JC, Levine M, Zeller RW (1997) Characterization of a notochord-specific enhancer from the Brachyury promoter region of the ascidian, Ciona intestinalis. Development 124(3):589–602
Corbo JC, Fujiwara S, Levine M, Di Gregorio A (1998) Suppressor of hairless activates brachyury expression in the Ciona embryo. Dev Biol 203(2):358–368
Dehal P, Satou Y, Campbell RK, Chapman J, Degnan B, De Tomaso A, Davidson B, Di Gregorio A et al (2002) The draft genome of Ciona intestinalis: insights into chordate and vertebrate origins. Science 298(5601):2157–2167
Delsuc F, Brinkmann H, Chourrout D, Philippe H (2006) Tunicates and not cephalochordates are the closest living relatives of vertebrates. Nature 439(7079):965–968
Denker E, Jiang D (2012) Ciona intestinalis notochord as a new model to investigate the cellular and molecular mechanisms of tubulogenesis. Semin Cell Dev Biol 23(3):308–319
Denker E, Bocina I, Jiang D (2013) Tubulogenesis in a simple cell cord requires the formation of bi-apical cells through two discrete par domains. Development 140(14):2985–2996
Denker E, Sehring IM, Dong B, Audisso J, Mathiesen B, Jiang D (2015) Regulation by a TGFβ-ROCK-actomyosin axis secures a non-linear lumen expansion that is essential for tubulogenesis. Development 142(9):1639–1650
Deschet K, Nakatani Y, Smith WC (2003) Generation of ci-Brachyury-GFP stable transgenic lines in the ascidian Ciona savignyi. Genesis 35(4):248–259
Di Gregorio A (2017) T-box genes and developmental gene regulatory networks in ascidians. Curr Top Dev Biol 122:55–91
Di Gregorio A, Levine M (1999) Regulation of Ci-tropomyosin-like, a Brachyury target gene in the ascidian, Ciona intestinalis. Development 126(24):5599–5609
Di Gregorio A, Corbo JC, Levine M (2001) The regulation of forkhead/HNF-3beta expression in the Ciona embryo. Dev Biol 229(1):31–43
Di Gregorio A, Harland RM, Levine M, Casey ES (2002) Tail morphogenesis in the ascidian, Ciona intestinalis, requires cooperation between notochord and muscle. Dev Biol 244(2):385–395
Dong B, Deng W, Jiang D (2011) Distinct cytoskeleton populations and extensive crosstalk control Ciona notochord tubulogenesis. Development 138(8):1631–1641
Dunn MP, Di Gregorio A (2009) The evolutionarily conserved leprecan gene: its regulation by Brachyury and its role in the developing Ciona notochord. Dev Biol 328(2):561–574
Evans AL, Faial T, Gilchrist MJ, Down T, Vallier L, Pedersen RA, Wardle FC, Smith JC (2012) Genomic targets of Brachyury (T) in differentiating mouse embryonic stem cells. PLoS One 7(3):e33346
Farley EK, Olson KM, Zhang W, Rokhsar DS, Levine MS (2016) Syntax compensates for poor binding sites to encode tissue specificity of developmental enhancers. Proc Natl Acad Sci U S A 113(23):6508–6513
Friedman JR, Kaestner KH (2006) The Foxa family of transcription factors in development and metabolism. Cell Mol Life Sci 63(19–20):2317–2328
Gluecksohn-Schoenheimer S (1940) The effect of an early lethal (t) in the house mouse. Genetics 25(4):391–400
Hikosaka A, Kusakabe T, Satoh N, Makabe KW (1992) Introduction and expression of recombinant genes in ascidian embryos. Develop Growth Differ 34:631–638
Hikosaka A, Kusakabe T, Satoh N (1994) Short upstream sequences associated with the muscle-specific expression of an actin gene in ascidian embryos. Dev Biol 166:763–769
Holland LZ, Laudet V, Schubert M (2004) The chordate amphioxus: an emerging model organism for developmental biology. Cell Mol Life Sci 61(18):2290–2308
Hotta K, Takahashi H, Erives A, Levine M, Satoh N (1999) Temporal expression patterns of 39 Brachyury-downstream genes associated with notochord formation in the Ciona intestinalis embryo. Develop Growth Differ 41(6):657–664
Hotta K, Takahashi H, Asakura T, Saitoh B, Takatori N, Satou Y, Satoh N (2000) Characterization of Brachyury-downstream notochord genes in the Ciona intestinalis embryo. Dev Biol 224(1):69–80
Hotta K, Mitsuhara K, Takahashi H, Inaba K, Oka K, Gojobori T, Ikeo K (2007a) A web-based interactive developmental table for the ascidian Ciona intestinalis, including 3D real-image embryo reconstructions: I. From fertilized egg to hatching larva. Dev Dyn 236(7):1790–1805
Hotta K, Yamada S, Ueno N, Satoh N, Takahashi H (2007b) Brachyury-downstream notochord genes and convergent extension in Ciona intestinalis embryos. Develop Growth Differ 49(5):373–382
Hotta K, Takahashi H, Satoh N, Gojobori T (2008) Brachyury-downstream gene sets in a chordate, Ciona intestinalis: integrating notochord specification, morphogenesis and chordate evolution. Evol Dev 10(1):37–51
Imai KS, Hino K, Yagi K, Satoh N, Satou Y (2004) Gene expression profiles of transcription factors and signaling molecules in the ascidian embryo: towards a comprehensive understanding of gene networks. Development 131(16):4047–4058
Imai KS, Levine M, Satoh N, Satou Y (2006) Regulatory blueprint for a chordate embryo. Science 312(5777):1183–1187
Irvine SQ (2013) Study of cis-regulatory elements in the Ascidian Ciona intestinalis. Curr Genomics 14(1):56–67
Jeffery WR, Ewing N, Machula J, Olsen CL, Swalla BJ (1998) Cytoskeletal actin genes function downstream of HNF-3beta in ascidian notochord development. Int J Dev Biol 42(8):1085–1092
Jeong Y, Epstein DJ (2003) Distinct regulators of Shh transcription in the floor plate and notochord indicate separate origins for these tissues in the mouse node. Development 130(16):3891–3902
Jiang D, Smith WC (2007) Ascidian notochord morphogenesis. Dev Dyn 236(7):1748–1757
Jiang D, Munro EM, Smith WC (2005) Ascidian prickle regulates both mediolateral and anterior-posterior cell polarity of notochord cells. Curr Biol 15(1):79–85
José-Edwards DS, Kerner P, Kugler JE, Deng W, Jiang D, Di Gregorio A (2011) The identification of transcription factors expressed in the notochord of Ciona intestinalis adds new potential players to the brachyury gene regulatory network. Dev Dyn 240(7):1793–1805
José-Edwards DS, Oda-Ishii I, Nibu Y, Di Gregorio A (2013) Tbx2/3 is an essential mediator within the Brachyury gene network during Ciona notochord development. Development 140(11):2422–2433
José-Edwards DS, Oda-Ishii I, Kugler JE, Passamaneck YJ, Katikala L, Nibu Y, Di Gregorio A (2015) Brachyury, Foxa2 and the cis-Regulatory Origins of the Notochord. PLoS Genet 11(12):e1005730
Katikala L, Aihara H, Passamaneck YJ, Gazdoiu S, José-Edwards DS, Kugler JE, Oda-Ishii I, Imai JH, Nibu Y, Di Gregorio A (2013) Functional Brachyury binding sites establish a temporal read-out of gene expression in the Ciona notochord. PLoS Biol 11(10):e1001697
Kispert A, Koschorz B, Herrmann BG (1995) The T protein encoded by Brachyury is a tissue-specific transcription factor. EMBO J 14(19):4763–4772
Kubo A, Suzuki N, Yuan X, Nakai K, Satoh N, Imai KS, Satou Y (2010) Genomic cis-regulatory networks in the early Ciona intestinalis embryo. Development 137(10):1613–1623
Kumano G, Yamaguchi S, Nishida H (2006) Overlapping expression of FoxA and Zic confers responsiveness to FGF signaling to specify notochord in ascidian embryos. Dev Biol 300(2):770–784
Lacalli T (2012) The Middle Cambrian fossil Pikaia and the evolution of chordate swimming. Evodevo 3(1):12
Lawson L, Harfe BD (2015) Notochord to nucleus pulposus transition. Curr Osteoporos Rep 13(5):336–341
Lemaire P (2009) Unfolding a chordate developmental program, one cell at a time: invariant cell lineages, short-range inductions and evolutionary plasticity in ascidians. Dev Biol 332(1):48–60
Levine M (2010) Transcriptional enhancers in animal development and evolution. Curr Biol 20(17):R754–R763
Liu GH, Mao CZ, Wu HY, Zhou DC, Xia JB, Kim SK, Cai DQ, Zhao H, Qi XF (2016) Expression profile of rrbp1 genes during embryonic development and in adult tissues of Xenopus laevis. Gene Expr Patterns 23–24:1–6
Mallatt J, Holland N (2013) Pikaia gracilens Walcott: stem chordate, or already specialized in the Cambrian? J Exp Zool B Mol Dev Evol 320:247–271
Matsumoto J, Kumano G, Nishida H (2007) Direct activation by Ets and Zic is required for initial expression of the Brachyury gene in the ascidian notochord. Dev Biol 306(2):870–882
Miyamoto DM, Crowther RJ (1985) Formation of the notochord in living ascidian embryos. J Embryol Exp Morphol 86:1–17
Morris SC, Caron JB (2012) Pikaia gracilens Walcott, a stem-group chordate from the middle Cambrian of British Columbia. Biol Rev Camb Philos Soc 87:480–512
Müller F, Chang B, Albert S, Fischer N, Tora L, Strähle U (1999) Intronic enhancers control expression of zebrafish sonic hedgehog in floor plate and notochord. Development 126(10):2103–2116
Myllyharju J, Kivirikko KI (1997) Characterization of the iron- and 2-oxoglutarate-binding sites of human prolyl 4-hydroxylase. EMBO J 16(6):1173–1180
Newman-Smith E, Kourakis MJ, Reeves W, Veeman M, Smith WC (2015) Reciprocal and dynamic polarization of planar cell polarity core components and myosin. Elife 13(4):e05361
Nibu Y, José-Edwards DS, Di Gregorio A (2013) From notochord formation to hereditary chordoma: the many roles of Brachyury. Biomed Res Int 2013:826435
Nishida H, Satoh N (1983) Cell lineage analysis in ascidian embryos by intracellular injection of a tracer enzyme. I. Up to the eight-cell stage. Dev Biol 99:382–394
Nishida H, Satoh N (1985) Cell lineage analysis in ascidian embryos by intracellular injection of a tracer enzyme. II. The 16- and 32-cell stages. Dev Biol 110:440–454
Nishiyama A, Fujiwara S (2008) RNA interference by expressing short hairpin RNA in the Ciona intestinalis embryo. Develop Growth Differ 50(6):521–529
Oda-Ishii I, Di Gregorio A (2007) Lineage-independent mosaic expression and regulation of the Ciona multidom gene in the ancestral notochord. Dev Dyn 236(7):1806–1819
Olsen CL, Jeffery WR (1997) A forkhead gene related to HNF-3beta is required for gastrulation and axis formation in the ascidian embryo. Development 124(18):3609–3619
Ortolani G (1954) Risultati definitive sulla distribuzione dei territory presuntivi degli organi nel germe di Ascidie allo stadio VIII, determinati con le marche al carbone. Pubbl Staz Zool Napoli 25:161–187
Passamaneck YJ, Di Gregorio A (2005) Ciona intestinalis: chordate development made simple. Dev Dyn 233(1):1–19
Passamaneck YJ, Katikala L, Perrone L, Dunn MP, Oda-Ishii I, Di Gregorio A (2009) Direct activation of a notochord cis-regulatory module by Brachyury and FoxA in the ascidian Ciona intestinalis. Development 136(21):3679–3689
Rastegar S, Hess I, Dickmeis T, Nicod JC, Ertzer R, Hadzhiev Y, Thies WG, Scherer G, Strähle U (2008) The words of the regulatory code are arranged in a variable manner in highly conserved enhancers. Dev Biol 318(2):366–377
Reese DE, Hall CE, Mikawa T (2004) Negative regulation of midline vascular development by the notochord. Dev Cell 6(5):699–708
Reverberi G (1971) Ascidians. In: Reverberi G (ed) Experimental embryology of marine and fresh-water invertebrates. North-Holland, Amsterdam, pp 507–550
Rhee JM, Oda-Ishii I, Passamaneck YJ, Hadjantonakis AK, Di Gregorio A (2005) Live imaging and morphometric analysis of embryonic development in the ascidian Ciona intestinalis. Genesis 43(3):136–147
Sasaki H, Hogan BL (1993) Differential expression of multiple fork head related genes during gastrulation and axial pattern formation in the mouse embryo. Development 118(1):47–59
Sasaki H, Yoshida K, Hozumi A, Sasakura Y (2014) CRISPR/Cas9-mediated gene knockout in the ascidian Ciona intestinalis. Develop Growth Differ 56(7):499–510
Sasakura Y, Suzuki MM, Hozumi A, Inaba K, Satoh N (2010) Maternal factor-mediated epigenetic gene silencing in the ascidian Ciona intestinalis. Mol Gen Genomics 283(1):99–110
Satoh N, Tagawa K, Takahashi H (2012) How was the notochord born? Evol Dev 14(1):56–75
Sawada A, Nishizaki Y, Sato H, Yada Y, Nakayama R et al (2005) Tead proteins activate the Foxa2 enhancer in the node in cooperation with a second factor. Development 132:4719–4729
Segade F, Cota C, Famiglietti A, Cha A, Davidson B (2016) Fibronectin contributes to notochord intercalation in the invertebrate chordate, Ciona intestinalis. EvoDevo 7(1):21
Sehring IM, Dong B, Denker E, Bhattachan P, Deng W, Mathiesen BT, Jiang D (2014) An equatorial contractile mechanism drives cell elongation but not cell division. PLoS Biol 12(2):e1001781
Sehring IM, Recho P, Denker E, Kourakis M, Mathiesen B, Hannezo E, Dong B, Jiang D (2015) Assembly and positioning of actomyosin rings by contractility and planar cell polarity. Elife 21(4):e09206
Smith J (1999) T-box genes: what they do and how they do it. Trends Genet 15(4):154–158
Søviknes AM, Glover JC (2008) Continued growth and cell proliferation into adulthood in the notochord of the appendicularian Oikopleura dioica. Biol Bull 214(1):17–28
Stemple DL (2005) Structure and function of the notochord: an essential organ for chordate development. Development 132:2503–2512
Stolfi A, Christiaen L (2012) Genetic and genomic toolbox of the chordate Ciona intestinalis. Genetics 192(1):55–66
Stolfi A, Gandhi S, Salek F, Christiaen L (2014) Tissue-specific genome editing in Ciona embryos by CRISPR/Cas9. Development 141(21):4115–4120
Tada M, Casey ES, Fairclough L, Smith JC (1998) Bix1, a direct target of Xenopus T-box genes, causes formation of ventral mesoderm and endoderm. Development 125(20):3997–4006
Takada N, Satoh N, Swalla BJ (2002) Expression of Tbx6, a muscle lineage T-box gene, in the tailless embryo of the ascidian Molgula tectiformis. Dev Genes Evol 212:354–356
Takahashi H, Hotta K, Erives A, Di Gregorio A, Zeller RW, Levine M, Satoh N (1999) Brachyury downstream notochord differentiation in the ascidian embryo. Genes Dev 13(12):1519–1523
Takahashi H, Hotta K, Takagi C, Ueno N, Satoh N, Shoguchi E (2010) Regulation of notochord-specific expression of Ci-Bra downstream genes in Ciona intestinalis embryos. Zool Sci 27(2):110–118
Tamplin OJ, Cox BJ, Rossant J (2011) Integrated microarray and ChIP analysis identifies multiple Foxa2 dependent target genes in the notochord. Dev Biol 360(2):415–425
Thompson JM, Di Gregorio A (2015) Insulin-like genes in ascidians: findings in Ciona and hypotheses on the evolutionary origins of the pancreas. Genesis 53(1):82–104
Urry LA, Whittaker CA, Duquette M, Lawler J, DeSimone DW (1998) Thrombospondins in early Xenopus embryos: dynamic patterns of expression suggest diverse roles in nervous system, notochord, and muscle development. Dev Dyn 211:390–407
Veeman MT, Nakatani Y, Hendrickson C, Ericson V, Lin C, Smith WC (2008) Chongmague reveals an essential role for laminin-mediated boundary formation in chordate convergence and extension movements. Development 135(1):33–41
Wada H, Okuyama M, Satoh N, Zhang S (2006) Molecular evolution of fibrillar collagen in chordates, with implications for the evolution of vertebrate skeletons and chordate phylogeny. Evol Dev 8:370–377
Weisblat DA, Sawyer RT, Stent GS (1978) Cell lineage analysis by intracellular injection of a tracer enzyme. Science 202:1295–1298
Yagi K, Satou Y, Satoh N (2004) A zinc finger transcription factor, ZicL, is a direct activator of Brachyury in the notochord specification of Ciona intestinalis. Development 131(6):1279–1288
Yamada S, Ueno N, Satoh N, Takahashi H (2011) Ciona intestinalis Noto4 contains a phosphotyrosine interaction domain and is involved in the midline intercalation of notochord cells. Int J Dev Biol 55(1):11–18
Yasuo H, Satoh N (1993) Function of vertebrate T gene. Nature 364(6438):582–583
Acknowledgements
Thanks to all present and past laboratory members and collaborators. We are particularly indebted to Drs. Diana José-Edwards, Lavanya Katikala, Izumi Oda-Ishii, and Yale Passamaneck for their original microphotographs. Research in our laboratory is supported by the National Institute of General Medical Sciences of the National Institutes of Health under award number R01GM100466.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2018 Springer Nature Singapore Pte Ltd.
About this chapter
Cite this chapter
Maguire, J.E., Pandey, A., Wu, Y., Di Gregorio, A. (2018). Investigating Evolutionarily Conserved Molecular Mechanisms Controlling Gene Expression in the Notochord. In: Sasakura, Y. (eds) Transgenic Ascidians . Advances in Experimental Medicine and Biology, vol 1029. Springer, Singapore. https://doi.org/10.1007/978-981-10-7545-2_8
Download citation
DOI: https://doi.org/10.1007/978-981-10-7545-2_8
Published:
Publisher Name: Springer, Singapore
Print ISBN: 978-981-10-7544-5
Online ISBN: 978-981-10-7545-2
eBook Packages: Biomedical and Life SciencesBiomedical and Life Sciences (R0)