Journal of Molecular Evolution

, Volume 71, Issue 1, pp 51–59 | Cite as

Evidence for the Heparin-Binding Ability of the Ascidian Xlink Domain and Insight into the Evolution of the Xlink Domain in Chordates

  • Masahiko Yoneda
  • Toshiya Nakamura
  • Miho Murai
  • Hiroshi Wada


The vertebrate Xlink domain is found in two types of genes: lecticans and their associated hyaluronan-and-proteoglycan-binding-link-proteins (HAPLNs), which are components of the extracellular matrix, and those represented by CD44 and stabilins, which are expressed on the surface of lymphocytes. In both types of genes, Xlink functions as a hyaluronan binding domain. We have already reported that protochordate ascidians possess only the latter type of gene. The present analysis of the expression of ascidian Xlink domain genes revealed that these genes function in blood cell migration and apoptosis. While the Xlink domain is found in various metazoans, including ascidians and nematodes, hyaluronan is believed to be specific for vertebrates. In comprehensive genome surveys for hyaluronan synthase (HAS), we found no HAS gene in ascidians. We also established that hyaluronan is absent from the ascidian body biochemically. Therefore, ascidians possess the Xlink domain, but they lack HA. We recovered one ascidian Xlink domain gene that encoded a heparin-binding protein, although it shows no affinity for hyaluronan. Based on these findings, we conclude that in invertebrates, the Xlink domain serves as heparin-binding protein domain and functions in blood cell migration and apoptosis. Its binding affinity for HA might have been acquired in the vertebrate lineage.


Hyaluronan Hyaluronan synthase (HAS) Ci-Link Xlink Ascidian Heparin 



We thank M. Ogasawara (Chiba University) for his valuable comment on the expression data. Ciona adults were provided by Y. Satou (Kyoto University) and M. Yoshida (University of Tokyo) through the National Bio-Resource Project of the MEXT, Japan. This study was supported by KAKENHI (Grant-in-Aid for Scientific Research) on Priority Areas ‘‘Comparative Genomics’’ from the Ministry of Education, Culture, Sports, Science and Technology of Japan to H.W.


  1. Abascal F, Zardoya R, Posada D (2005) ProtTest: selection of bestfit models of protein evolution. Bioinformatics 21:2104–2105CrossRefPubMedGoogle Scholar
  2. Banerji S (1999) LYVE-1, a new homologue of the CD44 glycoprotein is a lymph-specific receptor for hyaluronan. J Cell Biol 144:789–801CrossRefPubMedGoogle Scholar
  3. Bitter T, Muir HM (1962) A modified uronic acid carbazole reaction. Anal Biochem 4:330–334CrossRefPubMedGoogle Scholar
  4. Brissett NC, Perkins SJ (1996) The protein fold of the hyaluronate-binding proteoglycan tandem repeat domain of link protein, aggrecan and CD44 is similar to that of the C-type lectin superfamily. FEBS Lett 388:211–216CrossRefPubMedGoogle Scholar
  5. Burighel P, Cloney RA (1997) Urochordata: Ascidiacea. In: Harrison FW, Ruppert EE (eds) Microscopic anatomy of invertebrates. Wiley-Liss, New YorkGoogle Scholar
  6. Cavalcante MCM, Allodi S, Valente A-P, Straus AH, Takahashi HK, Mourão PAS, Pavão MSG (2000) Occurrence of heparin in the invertebrate Styela plicata (Tunicata) is restricted to cell layers facing the outside environment. J Biol Chem 275:36189–36196CrossRefPubMedGoogle Scholar
  7. Chambon J, Soule J, Pomies P, Fort P, Sahuquet A, Alexandre A, Mangeat P, Baghdiguian S (2002) Tail regression in Ciona intestinalis (Protochordate) involves a caspase-dependent apoptosis event associated with ERK activation. Development 129:3105–3114PubMedGoogle Scholar
  8. Cichy J, Pure E (2003) The liberation of CD44. J Cell Biol 161:839–843CrossRefPubMedGoogle Scholar
  9. Cima F, Basso G, Ballartin L (2003) Apoptosis and phosphatidylserine-mediated recoginition during the take-over phase of the colonial life-cycle in the ascidian Botryllus schlosseri. Cell Tissue Res 312:369–376CrossRefPubMedGoogle Scholar
  10. Day AJ, Prestwich G (2002) Hyaluronan-binding proteins: typing up the giant. J Biol Chem 277:4585–4588CrossRefPubMedGoogle Scholar
  11. Dehal P, Satou Y, Campbell PK et al (2002) The draft genome of Ciona intestinalis: insights into chordate and vertebrate origins. Science 298:2157–2167CrossRefPubMedGoogle Scholar
  12. Finn RD, Mistry J, Schuster-Böckler B, Griffiths-Jones S, Hollich V, Lassmann T, Moxon S, Marshall M, Khanna A, Durbin R, Eddy S, Sonnhammer ELL, Bateman A (2006) Pfam: clans, web tools and services. Nucleic Acids Res 34:D247–D251CrossRefPubMedGoogle Scholar
  13. Foster PA, Fulcher CA, Houghten RA, Zimmerman TS (1990) Synthetic factor VI11 peptides with amino acid sequences contained within the C2 domain of factor VI11 inhibit factor VI11 binding to phosphatidylserine. Blood 75:1999–2004PubMedGoogle Scholar
  14. Guindon S, Gascuel O (2003) A simple, fast and accurate algorithm to estimate large phylogenies by maximum likelihood. Syst Biol 52:696–704CrossRefPubMedGoogle Scholar
  15. Holt RA et al (2002) The genome sequence of the malaria mosquito Anopheles gambiae. Science 298:129–149CrossRefPubMedGoogle Scholar
  16. Kawashima T, Kawashima S, Tanaka C, Murai M, Yoneda M, Putnam NH, Rokhsar DS, Kanehisa M, Satoh N, Wada H (2009) Domain shuffling and the evolution of vertebrates. Genome Res 19:1393–1403CrossRefPubMedGoogle Scholar
  17. Kzhyshkowska J, Gratchev A, Goerdt S (2006) Stabilin-1, a homeostatic scavenger receptor with multiple functions. J Cell Mol Med 10:635–649CrossRefPubMedGoogle Scholar
  18. Lesley J, Gal I, Mahoney DJ, Cordell MR, Rugg MS, Hyman R, Day AJ, Mikecz K (2004) TSG-6 modulates the interaction between hyaluronan and cell surface CD44. J Biol Chem 279:25745–25754CrossRefPubMedGoogle Scholar
  19. Nottenburg C, Rees G, John TS (1989) Isolation of mouse CD44 cDNA: structural features are distinct from the primate cDNA. Proc Natl Acad Sci USA 86:8521–8525CrossRefPubMedGoogle Scholar
  20. Ogasawara M, Sasaki A, Metoki H, Shin-i T, Kohara Y, Satoh N, Satou Y (2002) Gene expression profiles in young adult Ciona intestinalis. Dev Genes Evol 212:173–185CrossRefPubMedGoogle Scholar
  21. Ohya T, Kaneko Y (1970) Novel hyaluronidase from streptomyces. Biochim Biophys Acta 198:607–609PubMedGoogle Scholar
  22. Ortel TL, Quinn-Allen MA, Keller FG, Peterson JA, Larocca D, Kane WH (1994) Localization of functionally important epitopes within the second C-type domain of coagulation factor V using recombinant chimeras. J Biol Chem 269:15898–15905PubMedGoogle Scholar
  23. Park S-Y, Kim S-Y, Jung M-Y, Bae D-J, Kim I-S (2008) Epidermal growth factor-like domain repeat of stabilin-2 recognizes phosphatidylserine during cell corpse clearance. Mol Cell Biol 28:5288–5298CrossRefPubMedGoogle Scholar
  24. Politz O, Gratchev A, McCourt PAG, Schledzewski K, Guillot P, Johansson S, Svineng G, Franke P, Kannicht C, Kzhyshkowska J, Longati P, Velten FW, Johansson S, Goerdt S (2002) Stabilin-1 and -2 constitute a novel family of fasciclin-like hyaluronan receptor homologues. Biochem J 362:155–164CrossRefPubMedGoogle Scholar
  25. Ponta H, Sherman L, Herrlich PA (2003) CD44: from adhesion molecules to signalling regulators. Nat Rev Mol Cell Biol 4:33–45CrossRefPubMedGoogle Scholar
  26. Prevo R, Banerji S, Ferguson DJP, Clasper S, Jackson DG (2001) Mouse LYVE-1 is an endocytic receptor for hyaluronan in lymphatic endothelium. J Biol Chem 276:19420–19430CrossRefPubMedGoogle Scholar
  27. Putnum N et al (2008) The amphioxus genome and the evolution of the chordate karyotype. Nature 453:1064–1072CrossRefGoogle Scholar
  28. Ruoslahti E (1996) Brain extracellular matrix. Glycobiol 6:489–492CrossRefGoogle Scholar
  29. Ruppert EE (1990) Structure, ultrastructure and function of the neural gland complex of Ascidia interrupta (Chordata, Ascidiacea): clarification of hypotheses regarding the evolution of the vertebrate pituitary. Acta Zool 71:135–149CrossRefGoogle Scholar
  30. Salzberg SL, White O, Peterson J, Eisen JA (2001) Microbial genes in the human genome: lateral transfer or gene loss. Science 292:1903–1906CrossRefPubMedGoogle Scholar
  31. Sandy JD, Flannery CR, Boynton RE, Neame PJ (1990) Isolation and characterization of disulfide-bonded peptides from the three globular domains of aggregating cartilage proteoglycan*. J Biol Chem 265:21108–21113PubMedGoogle Scholar
  32. Satou Y, Yamada L, Mochizuki Y, Takatori N, Kawashima T, Sasaki A, Hamaguchi M, Awazu S, Yagi K, Sasakura Y, Nakayama A, Ishikawa H, Inaba K, Satoh N (2002) A cDNA resource from the basal chordate Ciona intestinalis. Genesis 33:153–154CrossRefPubMedGoogle Scholar
  33. Satou Y, Kawashima T, Shoguchi E, Nakayama A, Satoh N (2005) An integrated database of the ascidian, Ciona intestinalis; Towards functional genomics. Zool Sci 22:837–843CrossRefPubMedGoogle Scholar
  34. Sea Urchin Genome Sequencing Consortium (2006) The genome of the sea urchin Strongylocentrotus purpuratus. Science 314:941–952CrossRefGoogle Scholar
  35. Small KS, Brundo M, Hill MM, Sidow A (2007) A haplotype alignment and reference sequence of the highly polymorphic Ciona savignyi genome. Genome Biol. 8:R41Google Scholar
  36. Spicer AP, Joo A, Bowling RA (2003) A hyaluronan binding link protein gene family whose members are physically linked adjacent to chondroitin sulfate proteoglycan core protein genes: the missing links. J Biol Chem 278:21083–21091CrossRefPubMedGoogle Scholar
  37. Tang LH, Rosenberg L, Reiner A, Poole AR (1979) Proteoglycans from bovine nasal cartilage. Properties of a soluble form of link protein. J Biol Chem 254:10523–10531PubMedGoogle Scholar
  38. Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG (1997) The ClustalX windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Res 24:4876–4882CrossRefGoogle Scholar
  39. Wada H (2010) Origin and genetic evolution of the vertebrate skeleton. Zool Sci 27:119–123CrossRefPubMedGoogle Scholar
  40. Weigel PH, Hascall VC, Tammi M (1997) Hyaluronan synthases. J Biol Chem 272:13997–14000CrossRefPubMedGoogle Scholar
  41. Wisniewski HG, Vilcek J (1996) TSG-6: an IL-I/TNF-inducible protein with anti-inflammatory activity. Cytokine Growth Factor Rev 8:143–156CrossRefGoogle Scholar
  42. Yamaguchi Y (2000) Lecticans: organizers of the brain extracellular matrix. Cell Mol Life Sci 57:276–289CrossRefPubMedGoogle Scholar
  43. Yasuo H, Satoh N (1994) An ascidian homolog of the mouse Brachyury (T) gene is expressed exclusively in notochord cells at fate restricted stage. Dev Growth Differ 36:9–18CrossRefGoogle Scholar
  44. Zhao J, Yoneda M, Takeyama M, Inoue Y, Kataoka T, Ohno-Jinno A, Isogai Z, Iwaki M, Zako M (2008) Competitive binding of heparin with hyaluronan to a specific motif in SPACR. J Neurochem 107:823–831CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Masahiko Yoneda
    • 1
  • Toshiya Nakamura
    • 2
  • Miho Murai
    • 1
  • Hiroshi Wada
    • 3
  1. 1.School of Nursing & HealthAichi Prefectural UniversityMoriyama-ku NagoyaJapan
  2. 2.Graduate School of Health SciencesHirosaki UniversityHirosakiJapan
  3. 3.Graduate School of Life and Environmental SciencesUniversity of TsukubaTsukubaJapan

Personalised recommendations