Skip to main content
Log in

The Mining of Pearl Formation Genes in Pearl Oyster Pinctada fucata by cDNA Suppression Subtractive Hybridization

  • Original Article
  • Published:
Marine Biotechnology Aims and scope Submit manuscript

Abstract

Recent researches revealed the regional preference of biomineralization gene transcription in the pearl oyster Pinctada fucata: it transcribed mainly the genes responsible for nacre secretion in mantle pallial, whereas the ones regulating calcite shells expressed in mantle edge. This study took use of this character and constructed the forward and reverse suppression subtractive hybridization (SSH) cDNA libraries. A total of 669 cDNA clones were sequenced and 360 expressed sequence tags (ESTs) greater than 100 bp were generated. Functional annotation associated 95 ESTs with specific functions, and 79 among them were identified from P. fucata at the first time. In the forward SSH cDNA library, it recognized mass amount of nacre protein genes, biomineralization genes dominantly expressed in the mantle pallial, calcium-ion-binding genes, and other biomineralization-related genes important for pearl formation. Real-time PCR showed that all the examined genes were distributed in oyster mantle tissues with a consistence to the SSH design. The detection of their RNA transcripts in pearl sac confirmed that the identified genes were certainly involved in pearl formation. Therefore, the data from this work will initiate a new round of pearl formation gene study and shed new insights into molluscan biomineralization.

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

Access this article

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

Similar content being viewed by others

References

  • Addadi L, Weiner S (1997) A pavement of pearl. Nature 389:912–915

    Article  CAS  Google Scholar 

  • Addadi L, Joester D, Nudelman F, Weiner S (2006) Mollusk shell formation: a source of new concepts for understanding biomineralization processes. Chem Eur J 12:980–987

    Article  PubMed  CAS  Google Scholar 

  • Che LM, Golubic S, Le Campion-Alsumard T, Payri C (2001) Developmental aspects of biomineralisation in the Polynesian pearl oyster Pinctada margaritifera var. cumingii. Oceanol Acta 24:S37–S49

    Article  Google Scholar 

  • Checa AG, Cartwright JHE, Willinger MG (2009) The key role of the surface membrane in why gastropod nacre grows in towers. Proc Natl Acad Sci U S A 106:38–43

    Article  PubMed  CAS  Google Scholar 

  • Diatchenko L, Lau YC, Campbell AP, Chenchik A, Moqadam F, Huang B, Lukyanov S, Lukyanov K, Gurskaya N, Sverdlov ED, Siebert PD (1996) Suppression subtractive hybridization: a method for generating differentially regulated or tissue-specific cDNA probes and libraries. Proc Natl Acad Sci U S A 93:6025–6030

    Article  PubMed  CAS  Google Scholar 

  • Diatchenko L, Lukyanov S, Lau YC, Siebert PD (1999) Suppression subtractive hybridization: a versatile method for identifying differentially expressed genes. Methods Enzymol 303:349–380

    Article  PubMed  CAS  Google Scholar 

  • Duplat D, Puissegur M, Bedouet L, Rousseau M, Boulzaguet H, Milet C, Sellos D, Wormhoudt AV, Lopez E (2006) Identification of calconectin, a calcium-binding protein specifically expressed by the mantle of Pinctada margaritifera. FEBS Lett 580:2435–2441

    Article  PubMed  CAS  Google Scholar 

  • Falini G, Albeck S, Weiner S, Addad L (1996) Control of aragonite or calcite polymorphism by mollusk shell macromolecules. Science 271:67–69

    Article  Google Scholar 

  • Gong N, Li Q, Huang J, Fang Z, Zhang G, Xie L, Zhang R (2008) Culture of outer epithelial cells from mantle tissue to study shell matrix protein secretion for biomineralization. Cell Tissue Res 333:493–501

    Article  PubMed  CAS  Google Scholar 

  • Hare PE (1963) Amino acids in the proteins from aragonite and calcite in the shells of Mytilus californianus. Science 139:216–217

    Article  PubMed  CAS  Google Scholar 

  • Huang X, Madan A (1999) CAP3: a DNA sequence assembly program. Genome Res 9:968–977

    Article  Google Scholar 

  • Huang J, Zhang C, Ma Z, Xie L, Zhang R (2007) A novel extracellular EF-hand protein involved in the shell formation of pearl oyster. Biochim Biophys Acta 1770:1037–1044

    Article  PubMed  CAS  Google Scholar 

  • Jackson DJ, McDougall C, Woodcroft B, Moase P, Rose RA, Kube M, Reinhardt R, Rokhsar DS, Montagnani C, Joubert C, Piquemal D, Degnan BM (2010) Parallel evolution of nacre building gene sites in molluscs. Mol Biol Evol 27:591–608

    Article  PubMed  CAS  Google Scholar 

  • Liu H, Liu S, Ge Y, Liu J, Wang X, Xie L, Zhang R, Wang Z (2007) Identification and characterization of a biomineralization related gene PFMG1 highly expressed in the mantle of Pinctada fucata. Biochemistry 46:844–851

    Article  PubMed  CAS  Google Scholar 

  • Ma Z, Huang J, Sun J, Wang G, Li C, Xie L, Zhang R (2007) A novel extrapallial fluid protein controls the morphology of nacre lamellae in the pearl oyster, Pinctada fucata. J Biol Chem 282:23253–23263

    Article  PubMed  CAS  Google Scholar 

  • Meyers MA, Lin AY, Chen P, Muyco J (2008) Mechanical strength of abalone nacre: role of the soft organic layer. J Mech Behav Biomed Mater 1:76–85

    Article  PubMed  Google Scholar 

  • Michenfelder M, Fu G, Lawrence G, Weaver JC, Wustman BA, Taranto L, Evans JS, Morse DE (2003) Characterization of two molluscan crystal-modulating biomineralization proteins and identification of putative mineral binding domains. Biopolymers 70:522–533

    Article  PubMed  CAS  Google Scholar 

  • Miyamoto H, Miyashita T, Okushima M, Nakano S, Morita T, Matsushiro A (1996) A carbonic anhydrase from the nacreous layer in oyster pearls. Proc Natl Acad Sci U S A 93:9657–9660

    Article  PubMed  CAS  Google Scholar 

  • Perrigault M, Tanguy A, Allam B (2009) Identification and expression of differentially expressed genes in the hard clam, Mercenaria mercenaria, in reponse to quahog parasite unknown (QPX). BMC Genomics 10:377

    Article  PubMed  Google Scholar 

  • Roberts S, Goetz G, White S, Goetz F (2009) Analysis of genes isolated from plated hemolytic of the Pacific oyster, Crassostreas gigas. Mar Biotechnol 11:24–44

    Article  PubMed  CAS  Google Scholar 

  • Rousseau M, Lopez E, Cute A, Mascara G, Smith DC, Maslin R, Borate X (2005) Sheet nacre growth mechanism: a Verona model. J Struct Biol 149:149–157

    Article  PubMed  Google Scholar 

  • Samata T, Hayashi N, Koon M, Hasegawa K, Hornito C, Akers S (1999) A new matrix protein family related to the nacreous layer formation of Pinctada fucata. FEBS Lett 462:225–229

    Article  PubMed  CAS  Google Scholar 

  • Sanchez S, Hordes S, Allier FH (2007) Identification of proteins involved in the functioning of Raffia pachyptila symbiosis by subtractive suppression hybridization. BMC Genomics 8:337

    Article  PubMed  Google Scholar 

  • Schefe JH, Lehmann KE, Buschmann IR, Unger T, Kaiser HF (2006) Quantitative real-time RT-PCR data analysis: current concepts and the novel “gene expression’s CT difference” formula. J Mol Med 84:901–910

    Article  PubMed  CAS  Google Scholar 

  • Shen X, Belcher AM, Hansma PK, Stucky GD, Morse DE (1997) Molecular cloning and characterization of lustrin A, a matrix protein from shell and pearl nacre of Haliotis rufescens. J Biol Chem 272:32472–32481

    Article  PubMed  CAS  Google Scholar 

  • Straub PF, Higham ML, Tanguy A, Landau BJ, Phoel WC, Hales LSJ, Thwing TK (2004) Suppression subtractive hybridization cDNA libraries to identify differentially expressed genes from contrasting fish habitats. Mar Biotechnology 6:386–399

    Article  CAS  Google Scholar 

  • Sudo S, Fujikawa T, Nagakura T, Ohkubo T, Sakaguchi K, Tanaka M, Nakashima K (1997) Structures of mollusc shell framework proteins. Nature 387:563–564

    Article  PubMed  CAS  Google Scholar 

  • Suzuki M, Saruwatari K, Kogure T, Yamamoto Y, Nishimura T, Kato T, Nagasawa H (2009) An acidic matrix protein, Pif, is a key macromolecule for nacre formation. Science 325:1388–1390

    Article  PubMed  CAS  Google Scholar 

  • Takeuchi T, Endo K (2006) Biphasic and dually coordinated expression of the genes encoding major shell matrix proteins in the pearl oyster Pinctada fucata. Mar Biotechnology 8:52–61

    Article  CAS  Google Scholar 

  • Takeuchi T, Sarashina I, Lijima M, Endo K (2008) In vitro regulation of CaCO3 crystal polymorphism by highly acidic molluscan shell protein Aspein. FEBS Lett 582:591–596

    Article  PubMed  CAS  Google Scholar 

  • Towe KM, Charles W, Harper J (1966) Pholidostrophiid Brachiopods: origin of the nacreous luster. Science 154:153–154

    Article  PubMed  CAS  Google Scholar 

  • Treccani L, Mann K, Heinemann F, Fritz M (2006) Perlwapin, an abalone nacre protein with three four-disulfide core (whey acidic protein) domains, inhibits the growth of calcium carbonate crystals. Biophys J 91:2601–2608

    Article  PubMed  CAS  Google Scholar 

  • Wang N, Kinoshita S, Riho C, Maeyama K, Nagai K, Watabe S (2009) Quantitative expression analysis of nacreous shell matrix protein genes in the process of pearl biogenesis. Comp Biochem Physiol B 154:346–350

    Article  PubMed  Google Scholar 

  • Xu W, Faisal M (2009) Identification of the molecules involved in zebra mussel (Dreissena polymorpha) hemolytic host defense. Comp Biochem Physiol B 154:143–149

    Article  PubMed  Google Scholar 

  • Yano M, Nagai K, Morimoto K, Miyamoto H (2006) Shematrin: a family of glycine-rich structural proteins in the shell of the pearl oyster Pinctada fucata. Comp Biochem Physiol B 144:254–262

    Article  PubMed  Google Scholar 

  • Zhang C, Zhang R (2006) Matrix proteins in the outer shells of molluscs. Mar Biotechnology 8:572–586

    Article  Google Scholar 

  • Zhang Y, Meng Q, Jiang T, Wang H, Xie L, Zhang R (2003) A novel ferritin subunit involved in shell formation from the pearl oyster (Pinctada fucata). Comp Biochem Physiol B 135:43–54

    Article  PubMed  Google Scholar 

  • Zhang C, Xie L, Huang J, Chen L, Zhang R (2006) A novel putative tyrosinase involved in periostracum formation from the pearl oyster (Pinctada fucata). Biochem Biophys Res Commun 342:632–639

    Article  PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Shugo Watabe.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Wang, N., Kinoshita, S., Nomura, N. et al. The Mining of Pearl Formation Genes in Pearl Oyster Pinctada fucata by cDNA Suppression Subtractive Hybridization. Mar Biotechnol 14, 177–188 (2012). https://doi.org/10.1007/s10126-011-9400-9

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10126-011-9400-9

Keywords

Navigation