Skip to main content
SpringerLink
Log in

Effects of cadmium exposure on sea urchin development assessed by SSH and RT-qPCR: metallothionein genes and their differential induction

  • Published:
Molecular Biology Reports Aims and scope Submit manuscript

Abstract

In order to study the defense strategies activated by Paracentrotus lividus embryos in response to sub-lethal doses of CdCl2, we compared the induced transcripts to that of control embryos by suppression subtractive hybridization technique. We isolated five metallothionein (MT) cDNAs and other genes related to detoxification, to signaling pathway components, to oxidative, reductive and conjugative biotransformation, to RNA maturation and protein synthesis. RT-qPCR analysis revealed that two of the five P. lividus MT (PlMT7 and PlMT8) genes appeared to be constitutively expressed and upregulated following cadmium treatment, whereas the other three genes (PlMT4, PlMT5, PlMT6) are specifically switched-on in response to cadmium treatment. Moreover, we found that this transcriptional induction is concentration dependent and that the cadmium concentration threshold for the gene activation is distinct for every gene. RT-qPCR experiments showed in fact that, among induced genes, PlMT5 gene is activated at a very low cadmium concentration (0.1 μM) whereas PlMT4 and PlMT6 are activated at intermediate doses (1–10 μM). Differently, PlMT7 and PlMT8 genes increase significantly their expression only in embryos treated with the highest dose (100 μM CdCl2). We found also that, in response to a lethal dose of cadmium (1 μM), only PlMT5 and PlMT6 mRNA levels increased further. These data suggest a hierarchical and orchestrated response of the P. lividus embryo to overcome differential environmental stressors that could interfere with a normal development.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

References

  1. Rainbow PS (2002) Kenneth Mellanby Review Award. Trace metal concentrations in aquatic invertebrates: why and so what? Environ Pollut 120:497–507

    Article  PubMed  CAS  Google Scholar 

  2. Viarengo A (1989) Heavy metal in marine invertebrates: mechanisms of regulation and toxicity at the cellular level. Aquat Sci 1:295–317

    CAS  Google Scholar 

  3. Wang Z, Yan C, Kong H, Wu D (2010) Mechanisms of cadmium toxicity to various trophic saltwater organisms. In: Parvau RG (ed) Cadmium in the environment. Nova Science Publishers Inc, New York, pp 297–336

    Google Scholar 

  4. Au DW, Lee CY, Chan KL, Wu RS (2001) Reproductive impairment of sea urchins upon chronic exposure to cadmium. Part I: effects on gamete quality. Environ Pollut 111:1–9

    Article  PubMed  CAS  Google Scholar 

  5. Matranga V, Toia G, Bonaventura R, Müller WE (2000) Cellular and biochemical responses to environmental and experimentally induced stress in sea urchin coelomocytes. Cell Stress Chaperones 5:113–120

    Article  PubMed  CAS  Google Scholar 

  6. Matranga V, Pinsino A, Celi M, Natoli A, Bonaventura R, Schröder HC, Müller WEG (2005) Monitoring chemical and physical stress using sea urchin immune cells. Prog Mol Subcell Biol 39:85–110

    Article  PubMed  CAS  Google Scholar 

  7. Radenac G, Fichet D, Miramand P (2001) Bioaccumulation and toxicity of four dissolved metals in Paracentrotus lividus sea-urchin embryo. Mar Environ Res 51:151–166

    Article  PubMed  CAS  Google Scholar 

  8. Roccheri MC, Matranga V (2010) Cellular, biochemical and molecular effects of cadmium on marine invertebrates: focus on Paracentrotus lividus sea urchin development. In: Parvau RG (ed) Cadmium in the environment. Nova Science Publishers Inc, New York, pp 337–366

    Google Scholar 

  9. Agnello M, Filosto S, Scudiero R, Rinaldi AM, Roccheri MC (2007) Cadmium induces an apoptotic response in sea urchin embryos. Cell Stress Chaperones 12:44–50

    Article  PubMed  CAS  Google Scholar 

  10. Filosto S, Roccheri MC, Bonaventura R, Matranga V (2008) Environmentally relevant cadmium concentrations affect development and induce apoptosis of Paracentrotus lividus larvae cultured in vitro. Cell Biol Toxicol 24:603–610. doi:10.1007/s10565-008-9066-x

    Article  PubMed  CAS  Google Scholar 

  11. Roccheri MC, Agnello M, Bonaventura R, Matranga V (2004) Cadmium induces the expression of specific stress proteins in sea urchin embryos. Biochem Biophys Res Commun 321:80–87. doi:10.1016/j.bbrc.2004.06.108

    Article  PubMed  CAS  Google Scholar 

  12. Russo R, Bonaventura R, Zito F, Schröder HC, Müller I, Müller WEG, Matranga V (2003) Stress to cadmium monitored by metallothionein gene induction in Paracentrotus lividus embryos. Cell Stress Chaperones 8:232–241

    Article  PubMed  CAS  Google Scholar 

  13. Schröder HC, Di Bella G, Janipour N, Bonaventura R, Russo R, Müller WEG, Matranga V (2005) DNA damage and developmental defects after exposure to UV and heavy metals in sea urchin cells and embryos compared to other invertebrates. Prog Mol Subcell Biol 39:111–137

    Article  PubMed  Google Scholar 

  14. Amiard J-C, Amiard-Triquet C, Barka S, Pellerin J, Rainbow PS (2006) Metallothioneins in aquatic invertebrates: their role in metal detoxification and their use as biomarkers. Aquat Toxicol 76:160–202. doi:10.1016/j.aquatox.2005.08.015

    Article  PubMed  CAS  Google Scholar 

  15. Chiarelli R, Agnello M, Roccheri MC (2011) Sea urchin embryos as a model system for studying autophagy induced by cadmium stress. Autophagy 7:1028–1034

    Article  PubMed  CAS  Google Scholar 

  16. Geraci F, Pinsino A, Turturici G, Savona R, Giudice G, Sconzo G (2004) Nickel, lead, and cadmium induce differential cellular responses in sea urchin embryos by activating the synthesis of different HSP70s. Biochem Biophys Res Commun 322:873–877. doi:10.1016/j.bbrc.2004.08.005

    Article  PubMed  CAS  Google Scholar 

  17. Nemer M, Wilkinson DG, Travaglini EC, Sternberg EJ, Butt TR (1985) Sea urchin metallothionein sequence: key to an evolutionary diversity. Proc Natl Acad Sci USA 82:4992–4994

    Article  PubMed  CAS  Google Scholar 

  18. Nemer M, Thornton RD, Stuebing EW, Harlow P (1991) Structure, spatial, and temporal expression of two sea urchin metallothionein genes, SpMTB1 and SpMTA. J Biol Chem 266:6586–6593

    PubMed  CAS  Google Scholar 

  19. Gianguzza F, Di Bernardo MG, Sollazzo M, Palla F, Ciaccio M, Carra E, Spinelli G (1989) DNA sequence and pattern of expression of the sea urchin (Paracentrotus lividus) alpha-tubulin genes. Mol Reprod Dev 1:170–181

    Article  PubMed  CAS  Google Scholar 

  20. Gianguzza F, Di Bernardo MG, Fais M, Palla F, Casano C, Russo R, Spinelli G (1990) Sequence and expression of Paracentrotus lividus alpha tubulin gene. Nucleic Acids Res 18:4915

    Article  PubMed  CAS  Google Scholar 

  21. Casano C, Ragusa M, Cutrera M, Costa S, Gianguzza F (1996) Spatial expression of alpha and beta tubulin genes in the late embryogenesis of the sea urchin Paracentrotus lividus. Int J Dev Biol 40:1033–1041

    PubMed  CAS  Google Scholar 

  22. Casano C, Roccheri MC, Onorato K, Cascino D, Gianguzza F (1998) Deciliation: a stressful event for Paracentrotus lividus embryos. Biochem Biophys Res Commun 248:628–634

    Article  PubMed  CAS  Google Scholar 

  23. Scudiero R, Capasso C, Del Vecchio-Blanco F, Savino G, Capasso A, Parente A, Parisi E (1995) Isolation and primary structure determination of a metallothionein from Paracentrotus lividus (Echinodermata, Echinoidea). Comp Biochem Physiol B 111:329–336

    Article  PubMed  CAS  Google Scholar 

  24. Binz PA, Kägi JHR (1999) Metallothionein: molecular evolution and classification. In: Klaassen C (ed) Metallothionein IV. Birkhäuser Verlag, Basel, pp 7–13

    Chapter  Google Scholar 

  25. Capdevila M, Atrian S (2011) Metallothionein protein evolution: a miniassay. J Biol Inorg Chem 16:977–989. doi:10.1007/s00775-011-0798-3

    Article  PubMed  CAS  Google Scholar 

  26. de Torres M, Sanchez P, Fernandez-Delmond I, Grant M (2003) Expression profiling of the host response to bacterial infection: the transition from basal to induced defence responses in RPM1-mediated resistance. Plant J 33:665–676

    Article  PubMed  Google Scholar 

  27. Goldstone JV, Hamdoun A, Cole BJ, Howard-Ashby M, Nebert DW, Scally M, Dean M, Epel D, Hahn ME, Stegeman JJ (2006) The chemical defensome: environmental sensing and response genes in the Strongylocentrotus purpuratus genome. Dev Biol 300:366–384. doi:10.1016/j.ydbio.2006.08.066

    Article  PubMed  CAS  Google Scholar 

  28. Goldstone JV (2008) Environmental sensing and response genes in cnidaria: the chemical defensome in the sea anemone Nematostella vectensis. Cell Biol Toxicol 24:483–502. doi:10.1007/s10565-008-9107-5

    Article  PubMed  CAS  Google Scholar 

  29. Epel D (2003) Protection of DNA during early development: adaptations and evolutionary consequences. Evol Dev 5:83–88

    Article  PubMed  CAS  Google Scholar 

  30. Hamdoun A, Epel D (2007) Embryo stability and vulnerability in an always changing world. Proc Natl Acad Sci USA 104(6):1745–1750. doi:10.1073/pnas.0610108104

    Article  PubMed  CAS  Google Scholar 

  31. Damle SS, Davidson EH (2012) Synthetic in vivo validation of gene network circuitry. Proc Natl Acad Sci USA 109(5):1548–1553. doi:10.1073/pnas.1119905109

    Article  PubMed  CAS  Google Scholar 

  32. Moncada S, Erusalimsky JD (2002) Does nitric oxide modulate mitochondrial energy generation and apoptosis? Nat Rev Mol Cell Biol 3:214–220. doi:10.1038/nrm762

    Article  PubMed  CAS  Google Scholar 

  33. Xu W, Charles IG, Moncada S (2005) Nitric oxide: orchestrating hypoxia regulation through mitochondrial respiration and the endoplasmic reticulum stress response. Cell Res 15:63–65. doi:10.1038/sj.cr.7290267

    Article  PubMed  Google Scholar 

  34. Angerer LM, Kawczynski G, Wilkinson DG, Nemer M, Angerer RC (1986) Spatial patterns of metallothionein mRNA expression in the sea urchin embryo. Dev Biol 116:543–547

    Article  PubMed  CAS  Google Scholar 

  35. Vasák M, Hasler DW (2000) Metallothioneins: new functional and structural insights. Curr Opin Chem Biol 4:177–183

    Article  PubMed  Google Scholar 

  36. Wilkinson DG, Nemer M (1987) Metallothionein genes MTa and MTb expressed under distinct quantitative and tissue-specific regulation in sea urchin embryos. Mol Cell Biol 7:48–58

    PubMed  CAS  Google Scholar 

  37. Capdevila M, Bofill R, Palacios Ò, Atrian S (2012) State-of-the-art of metallothioneins at the beginning of the 21st century. Coord Chem Rev 256:46–62. doi:10.1016/j.ccr.2011.07.006

    Article  CAS  Google Scholar 

  38. Guirola M, Naranjo Y, Capdevila M, Atrian S (2011) Comparative genomics analysis of metallothioneins in twelve Drosophila species. J Inorg Biochem 105:1050–1059. doi:10.1016/j.jinorgbio.2011.05.004

    Article  PubMed  CAS  Google Scholar 

  39. Höckner M, Stefanon K, de Vaufleury A, Monteiro F, Pérez-Rafael S, Palacios O, Capdevila M, Atrian S, Dallinger R (2011) Physiological relevance and contribution to metal balance of specific and non-specific Metallothionein isoforms in the garden snail Cantareus aspersus. Biometals 24:1079–1092. doi:10.1007/s10534-011-9466-x

    Article  PubMed  Google Scholar 

  40. Thirumoorthy N, Shyam Sunder A, Manisenthil Kumar K, Senthil Kumar M, Ganesh G, Chatterjee M (2011) A review of metallothionein isoforms and their role in pathophysiology. World J Surg Oncol 9:54. doi:10.1186/1477-7819-9-54

    Article  PubMed  CAS  Google Scholar 

  41. Soazig L, Marc L (2003) Potential use of the levels of the mRNA of a specific metallothionein isoform (MT-20) in mussel (Mytilus edulis) as a biomarker of cadmium contamination. Mar Pollut Bull 46:1450–1455. doi:10.1016/S0025-326X(03)00283-2

    Article  PubMed  CAS  Google Scholar 

  42. Wang W-X, Rainbow PS (2010) Significance of metallothioneins in metal accumulation kinetics in marine animals. Comp Biochem Physiol C 152:1–8. doi:10.1016/j.cbpc.2010.02.015

    Google Scholar 

  43. Zeitoun-Ghandour S, Charnock JM, Hodson ME, Leszczyszyn OI, Blindauer CA, Stürzenbaum SR (2010) The two Caenorhabditis elegans metallothioneins (CeMT-1 and CeMT-2) discriminate between essential zinc and toxic cadmium. FEBS J 277(11):2531–2542. doi:10.1111/j.1742-4658.2010.07667.x

    Article  PubMed  CAS  Google Scholar 

  44. Mao H, Wang D-H, Yang W-X (2012) The involvement of metallothionein in the development of aquatic invertebrate. Aquat Toxicol 110–111:208–213. doi:10.1016/j.aquatox.2012.01.018

    Article  PubMed  Google Scholar 

Download references

Acknowledgments

We would like to thank C. Luparello and V. Matranga for their critical reading and feedbacks on this manuscript. We would also like to apologize with all our colleagues whose work was not properly cited due to space restriction. This work was supported by MIUR (ex 60 %) grant to F.G.

Author information

Authors and Affiliations

  1. Dipartimento di Scienze e Tecnologie Molecolari e Biomolecolari, Università di Palermo, viale delle Scienze, Ed. 16, 90128, Palermo, Italy

    Maria Antonietta Ragusa, Salvatore Costa, Marco Gianguzza, Maria Carmela Roccheri & Fabrizio Gianguzza

Authors
  1. Maria Antonietta Ragusa
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Salvatore Costa
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Marco Gianguzza
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Maria Carmela Roccheri
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Fabrizio Gianguzza
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Maria Antonietta Ragusa.

Electronic supplementary material

Rights and permissions

Reprints and permissions

About this article

Cite this article

Ragusa, M.A., Costa, S., Gianguzza, M. et al. Effects of cadmium exposure on sea urchin development assessed by SSH and RT-qPCR: metallothionein genes and their differential induction. Mol Biol Rep 40, 2157–2167 (2013). https://doi.org/10.1007/s11033-012-2275-7

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11033-012-2275-7

Keywords

Search

Navigation