Roux's archives of developmental biology

, Volume 203, Issue 6, pp 320–327 | Cite as

Expression of AMD 1, a gene for a MyoD 1-related factor in the ascidian Halocynthia roretzi

  • sato Araki
  • Hidetoshi Saiga
  • Kazuhiro W. Makabe
  • Noriyuki Satoh
Original Articles


We have isolated and cloned a gene, designated AMD1 (ascidian MyoD-related factor 1), and its cDNAs that encode a member of the family of myogenic basic helix-loop-helix (bHLH) factors from the ascidian Halocynthia roretzi. The AMD1 gene consists of four exons and is transcribed into at least two distinct mRNAs, which differ in their 3' untranslated region. The gene encodes a protein of 435 amino acids, which exhibits homology to the bHLH domain of other myogenic bHLH factors including vertebrate MyoDt. A reverse transcription-polymerase chain reaction (RT-PCR) assay revealed that transcripts of the gene were not detectable in fertilized eggs or in very early embryos. They were first detected at the 64-cell stage, a few hours prior to the accumulation of mRNAs for embryonic muscle-specific proteins. In addition, expression of the AMD1 gene was evident in adult body-wall muscle but not in heart and other nonmuscle tissues. These results suggest the possibility that, in ascidians as in vertebrates, the myogenic bHLH factor is involved in the specification of embryonic cells as myogenic cells.

Key words

Ascidian embryos Myogenesis Helixloop-helix protein Body-wall muscle 


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  1. Benezra R, Davis RL, Lockshon D, Turner DL, Weintraub H (1990) The protein Id: A negative regulator of helix-loop-helix DNA binding proteins. Cell 61:49–59Google Scholar
  2. Bengal E, Ransone L, Scharfmann R, Dwarki VJ, Tapscott SJ, Weintraub H, Verma IM (1992) Functional antagonism between c-Jun and MyoD proteins: A direct physical association. Cell 68:507–519Google Scholar
  3. Braun T, Buschhausen-Denker G, Bober E, Tannich E, Arnold HH (1989) A novel human muscle factor related to but distinct from MyoD1 induces myogenic conversion in 10T1/2 fibroblasts. EMBO J 8:701–709Google Scholar
  4. Brennan TJ, Chakraborty T, Olson EN (1991) Mutagenesis of the myogenin basic region identifies an ancient protein motif critical for activation of myogenesis. Proc Natl Acad Sci USA 88:5675–5679Google Scholar
  5. Chen L, Krause M, Draper B, Weintraub H, Fire A (1992) Bodywall muscle formation in Caenorhabditis elegans embryos that lack the MyoD homolog hlh-1. Science 256:240–243Google Scholar
  6. Chomczynski P, Sacchi N (1987) Single-step method of RNA isolation by acid guanidium thiocyanate-phenol-chloroform extraction. Anal Biochem 162:156–159Google Scholar
  7. Clark TG, Morris J, Akamatsu M, McGraw RA, Ivarie R (1990) A bovine homolog to the human myogenic determination factor myf-5: Sequence conservation and 3′ processing of transcripts. Nucleic Acids Res 18:3147–3153Google Scholar
  8. Conklin EG (1905) The organization and cell lineage of the ascidian egg. J Acad Natl Sci Philadelphia 13:1–119Google Scholar
  9. Davis RL, Weintraub H (1992) Acquisition of myogenic specificity by replacement of three amino acid residues from MyoD into E12. Science 256:1027–1030Google Scholar
  10. Davis RL, Weintraub H, Lassar AB (1987) Expression of a single transfected cDNA converts fibroblasts to myoblasts. Cell 51:987–1000Google Scholar
  11. Fujisawa-Sehara A, Nabeshima Y, Hosoda Y, Obinata T, Nabeshima Y (1990) Myogenin contains two domains conserved among myogenic factors. J Biol Chem 265:15219–15223Google Scholar
  12. Hasty P, Bradley A, Morris JH, Edmondson DG, Venuti JM, Olson EN, Klein WH (1993) Muscle deficiency and neonatal death in mice with a targeted mutation in the myogenin gene. Nature 364:501–506Google Scholar
  13. Hikosaka A, Kusakabe T, Satoh N, Makabe KW (1992) Introduction and expression of recombinant genes in ascidian embryos. Dev Growth Differ 34:627–634Google Scholar
  14. Hikosaka A, Satoh N, Makabe KW (1993) Regulated spatial expression of fusion gene constructs with the 5′-upstream region of Halocynthia roretzi muscle actin gene in Ciona savignyi embryos. Roux's Arch Dev Biol, in pressGoogle Scholar
  15. Hinterberger TJ, Mays JL, Konieczny SF (1992) Structure and myofiber-specific expression of the rat muscle regulatory gene MRF4. Gene 117:201–207Google Scholar
  16. Hopwood ND, Pluck A, Gurdon JB (1989) MyoDl expression in the forming somites is an early response to mesoderm induction in Xenopus embryos. EMBO J 8:3409–3417Google Scholar
  17. Jen Y, Weintraub H, Benezra R (1992) Overexpression of Id protein inhibits the muscle differentiation program: In vivo association of Id with E2A proteins. Genes Dev 6:1466–1479Google Scholar
  18. Krause M, Fire A, Harrison SW, Priess J, Weintraub H (1990) CeMyoD accumulation defines the body wall muscle cell fate during C. elegans embryogenesis. Cell 63:907–919Google Scholar
  19. Kusakabe T, Suzuki J, Saiga H, Jeffery WR, Makabe KW, Satoh N (1991) Temporal and spatial expression of a muscle actin gene during embryogenesis of the ascidian Halocynthia roretzi. Dev Growth Differ 33:227–234Google Scholar
  20. Kusakabe T, Makabe KW, Satoh N (1992) Tunicate muscle actin genes. Structure and organization as a gene cluster. J Mol Biol 227:955–960Google Scholar
  21. Lassar AB, Buskin JN, Lockshon D, Davis RL, Apone S, Hauschka SD, Weintraub H (1989) MyoD1 is a sequence-specific DNA binding protein requiring a region of myc homology to bind to the muscle creatine kinase enhancer. Cell 58:823–831Google Scholar
  22. Li L, Chambard J-C, Karin M, Olson EN (1992a) Fos and Jun repress transcriptional activation by myogenin and MyoD: The amino terminus of Jun can mediate repression. Genes Dev 6:676–689Google Scholar
  23. Li L, Zhou J, James G, Heller-Harrison R, Czech MP, Olson EN (1992b) FGF inactivates myogenic helix-loop-helix proteins through phosphorylation of a conserved protein kinase C site in their DNA-binding domain. Cell 71:1181–1194Google Scholar
  24. Lin Z, Dechesne CA, Eldridge J, Paterson BM (1989) An avian muscle factor related to MyoDl activates muscle-specific promoters in nonmuscle cells of different germ-layer origin and in BrdU-treated myoblasts. Genes Dev 3:986–996Google Scholar
  25. Makabe KW, Fujiwara S, Saiga H, Satoh N (1990) Specific expression of myosin heavy chain gene in muscle lineage cells of the ascidian embryo. Roux's Arch Dev Biol 199:307–313Google Scholar
  26. Michelson AM, Abmayr SM, Bate M, Arias AM, Maniatis T (1990) Expression of a MyoD family member prefigures muscle pattern in Drosophila embryos. Genes Dev 4:2086–2097Google Scholar
  27. Miner JH, Wold B (1990) Herculin, a fourth member of the MyoD family of myogenic regulatory genes. Proc Natl Acad Sci USA 87:1089–1093Google Scholar
  28. Murre C, McCaw PS, Vaessin H, Candy M, Jan LY, Jan YN, Cabrera CV, Buskin JN, Hauschka SD, Lassar AB, Weintraub H, Baltimore D (1989) Interactions between heterologous helixloop-helix proteins generate complexes that bind specifically to a common DNA sequence. Cell 58:537–544Google Scholar
  29. Nishida H (1987) Cell lineage analysis in ascidian embryos by intracellular injection of a tracer enzyme. III. Up to the tissue restricted stage. Dev Biol 121:526–541Google Scholar
  30. Ott M-O, Bober E, Lyons G, Arnold H, Buckingham M (1991) Early expression of the myogenic regulatory gene, myf-5, in precursor cells of skeletal muscle in the mouse embryo. Development 111:1097–1107Google Scholar
  31. Paterson BM, Walldorf U, Eldridge J, Dubendorfer A, Frasch M, Gehring WJ (1991) The Drosophila homologue of vertebrate myogenic-determination genes encodes a transiently expressed nuclear protein marking primary myogenic cells. Proc Natl Acad Sci USA 88:3782–3786Google Scholar
  32. Pownall ME, Emerson Jr CP (1992) Sequential activation of three myogenic regulatory genes during somite morphogenesis in quail embryos. Dev Biol 151:67–79Google Scholar
  33. Rhodes SJ, Konieczny SF (1989) Identification of MRF4: A new member of the muscle regulatory factor gene family. Genes Dev 3:2050–2061Google Scholar
  34. Saiki RK, Gelfand DH, Stoffel S, Scharf SJ, Higuchi R, Horn GT, Mullis KB, Erlich HA (1987) Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science 239:487–491Google Scholar
  35. Salminen A, Braun T, Buchberger A, Jurs S, Winter B, Arnold H-H (1991) Transcription of the muscle regulatory gene MYF-4 is regulated by serum components, peptide growth factors and signaling pathways involving G proteins. J Cell Biol 115:905–917Google Scholar
  36. Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: A laboratory manual, 2nd edn. Cold Spring Harbor Laboratory Press, New YorkGoogle Scholar
  37. Satoh N (1979) On the ‘clock’ mechanism determining the time of tissue-specific enzyme development during ascidian embryogenesis. I. Acetylcholinesterase development in cleavage-arrested embryos. J Embryol Exp Morphol 54:131–139Google Scholar
  38. Satoh N (1993) Developmental biology of ascidians. Cambridge Univ Press, Cambridge New York (in press)Google Scholar
  39. Satoh N, Deno T, Nishida H, Nishikata T, Makabe KW (1990) Cellular and molecular mechanisms of muscle cell differentiation in ascidian embryos. Int Rev Cytol 122:221–258Google Scholar
  40. Vaidya TB, Rhodes SJ, Moore JL, Sherman DA, Konieczny SF, Taparowsky EJ (1992) Isolation and structural analysis of the rat MyoD gene. Gene 116:223–230Google Scholar
  41. Venuti JM, Jeffery WR (1989) Cell lineage and determination of cell fate in ascidian embryos. Int J Dev Biol 33:197–212Google Scholar
  42. Venuti JM, Goldberg L, Chakraborty T, Olson EN, Klein WH (1991) A myogenic factor from sea urchin embryos capable of programming muscle differentiation in mammalian cells. Proc Natl Acad Sci USA 88:6219–6223Google Scholar
  43. Weintraub H, Tapscott SJ, Davis RL, Thayer MJ, Adam MA, Lassar AB, Miller AD (1989) Activation of muscle-specific genes in pigment, nerve, fat, liver, and fibroblast cell lines by forced expression of MyoD. Proc Natl Acad Sci USA 86:5434–5438Google Scholar
  44. Weintraub H, Dwarki VJ, Verma I, Davis R, Hollenberg S, Snider L, Lassar A, Tapscott SJ (1991) Muscle-specific transcriptional activation by MyoD Genes Dev 5:1377–1386Google Scholar
  45. Wright WE, Sassoon DA, Lin VK (1989) Myogenin, a factor regulating myogenesis, has a domain homologous to MyoD. Cell 56:607–617Google Scholar

Copyright information

© Springer-Verlag 1994

Authors and Affiliations

  • sato Araki
    • 1
  • Hidetoshi Saiga
    • 2
  • Kazuhiro W. Makabe
    • 1
  • Noriyuki Satoh
    • 1
  1. 1.Department of Zoology, Faculty of ScienceKyoto UniversityKyotoJapan
  2. 2.Department of Biology, Faculty of ScienceTokyo Metropolitan UniversityHachi-ojiJapan

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