Abstract
Molluscan shells are composed of calcium carbonates, with small amounts of extracellular matrices secreted from mantle epithelial cells. Many types of shell matrix proteins (SMPs) have been identified from molluscan shells or mantle cells. The pen shell Atrina pectinata (Pinnidae) has two different shell microstructures, the nacreous and prismatic layers. Nacreous and prismatic layer-specific matrix proteins have been reported in Pteriidae bivalves, but remain unclear in Pinnidae. We performed transcriptome analysis using the mantle cells of A. pectinata to screen the candidate transcripts involved in its prismatic layer formation. We found Asprich and nine highly conserved prismatic layer-specific SMPs encoding transcript in P. fucata, P. margaritifera, and P. maxima (Tyrosinase, Chitinase, EGF-like proteins, Fibronectin, valine-rich proteins, and prismatic uncharacterized shell protein 2 [PUSP2]) using molecular phylogenetic analysis or multiple alignment. We confirmed these genes were expressed in the epithelial cells of the mantle edge (outer surface of the outer fold) and the mantle pallium. Phylogenetic character mapping of these SMPs was used to infer a possible evolutionary scenario of them in Pteriomorphia. EGF-like proteins, Fibronectin, and valine-rich proteins encoding genes each evolved in the linage leading to four Pteriomorphia (Mytilidae, Pinnidae, Ostreidae, and Pteriidae), PUSP2 evolved in the linage leading to three Pteriomorphia families (Pinnidae, Ostreidae, and Pteriidae), and chitinase was independently evolved as SMPs in Mytilidae and in other Pteriomorphia (Pinnidae, Ostreidae, and Pteriidae). Our results provide a new dataset for A. pectinata SMP annotation, and a basis for understanding the evolution of prismatic layer formation in bivalves.
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Data Availability
Transcriptome data are available in the DNA Data Bank of Japan (DDBJ) (Sequence Read Archive: DRR209159, BioProject: PRJDB9333, BioSample: SAMD00206368, TSA: ICPQ01000001-ICPQ01061263). Other sequence data (ApeAsprich, ApeCR-CN1, ApeCR-CN2, ApeEGF-like 1, ApeEGF-like 2, ApeCR-FN, ApeMP10, ApeAlveolin, and ApePUSP2) are available in the DDBJ under accession numbers LC574994-LC575002.
References
Aguilera F, McDougall C, Degnan BM (2014) Evolution of the tyrosinase gene family in bivalve molluscs: independent expansion of the mantle gene repertoire. Acta Biomater 10(9):3855–3865. https://doi.org/10.1016/j.actbio.2014.03.031
Amphlett GW, Hrinda ME (1983) The binding of calcium to human fibronectin. Biochem Biophys Res Commun 111:1045–1053. https://doi.org/10.1016/0006-291x(83)91405-5
Belcher A, Wu X, Christensen R et al (1996) Control of crystal phase switching and orientation by soluble mollusc-shell proteins. Nature 381:56–58. https://doi.org/10.1038/381056a0
Capella-Gutiérrez S, Silla-Martínez JM, Gabaldón T (2009) trimAl: a tool for automated alignment trimming in large-scale phylogenetic analyses. Bioinformatics (Oxford, England) 25:1972–1973. https://doi.org/10.1093/bioinformatics/btp348
Davison NM, Oshlack A (2014) Corset: enabling differential gene expression analysis for de novoassembled transcriptomes. Genome Biol 15:410. https://doi.org/10.1186/s13059-014-0410-6
Feng D, Li Q, Yu H, Kong L, Du S (2017) Identification of conserved proteins from diverse shell matrix proteome in Crassostrea gigas: characterization of genetic bases regulating shell formation. Sci Rep 7:45754. https://doi.org/10.1038/srep45754
Finn RD, Coggill P, Eberhardt RY et al (2016) The Pfam protein families database: towards a more sustainable future. Nucleic Acids Res 44:D279–D285. https://doi.org/10.1093/nar/gkv1344
Gao P, Liao Z, Wang XX, Bao LF, Fan MH, et al (2015) Layer-by-layer proteomic analysis of mytilus galloprovincialis shell. PLoS One 10(7):e0133913. https://doi.org/10.1371/journal.pone
Gotliv B, Kessler N, Sumerel JL et al (2005) Asprich: a novel aspartic acid-rich protein family from the prismatic shell matrix of the bibalve Atrina rigida. Chem Bio Chem 6:304–314. https://doi.org/10.1002/cbic.200400221
Götz S, García-Gómez JM, Terol J, Williams TD, Nagaraj SH, et al (2008) High-throughput functional annotation and data mining with the Blast2GO suite. Nucleic Acids Res 36(10):3420–35. https://doi.org/10.1093/nar/gkn176
Grabherr MG, Haas BJ, Yassour M et al (2011) Full-length transcriptome assembly from RNA-Seq data without a reference genome. Nat Biotechnol 29:644–652. https://doi.org/10.1038/nbt.1883
Haas BJ, Papanicolaou A, Yassour M et al (2013) De novo transcript sequence reconstruction from RNA-seq using the Trinity platform for reference generation and analysis. Nat Protoc 8:1494–1512. https://doi.org/10.1038/nprot.2013.084
Iseli C, Jongeneel CV, Bucher P (1999) ESTScan: a program for detecting, evaluating, and reconstructing potential coding regions in EST sequences. Proc Int Conf Intell Syst Mol Biol 138–48.
Isowa Y, Sarashina I, Setiamarga DHE, Endo K (2012) A comparative study of the shell matrix protein aspein in pterioid bivalves. J Mol Evol 75:11–18. https://doi.org/10.1007/s00239-012-9514-3
Iwamoto S, Shimizu K, Negishi L, Suzuki N, Nagata K, Suzuki M (2020) Characterization of the chalky layer-derived EGF-like domain-containing protein (CgELC) in the pacific oyster, Crassostrea gigas. J Struct Biol 212(1):107594. https://doi.org/10.1016/j.jsb.2020.107594
Jackson DJ, McDougall C, Green K et al (2006) A rapidly evolving secretome builds and patterns a sea shell. BMC Biol 4:40. https://doi.org/10.1186/1741-7007-4-40
Jackson DJ, McDougall C, Woodcroft B et al (2010) Parallel evolution of nacre building gene sets in molluscs. Mol Biol Evol 27:591–608. https://doi.org/10.1093/molbev/msp278
Jackson DJ, Mann K, Häussermann V et al (2015) The Magellania venosa biomineralizing proteome: a window into brachiopod shell evolution. Gen Biol Evol 7:1349–1362. https://doi.org/10.1093/gbe/evv074
Liu C, Li S, Kong J, Liu Y, Wang T, Xie L, Zhang R (2015) In-depth proteomic analysis of shell matrix proteins of Pinctada fucata. Sci Rep 5:17269. https://doi.org/10.1093/gbe/evv074
Katoh K, Misawa K, Ki K, Miyata T (2002) MAFFT: a novel method for rapid multiple sequence alignment based on fast Fourier transform. Nucleic Acids Res 30:3059–3066. https://doi.org/10.1093/nar/gkf436
Khamchun S, Sueksakit K, Chaiyarit S, Thongboonkerd V (2019) Modulatory effects of fibronectin on calcium oxalate crystallization, growth, aggregation, adhesion on renal tubular cells, and invasion through extracellular matrix. J Biol Inorg Chem 24:235–246. https://doi.org/10.1007/s00775-019-01641-w
Kintsu H, Okumura T, Negishi L et al (2017) Crystal defects induced by chitin and chitinolytic enzymes in the prismatic layer of Pinctada fucata. Biochem Biophys Res Commun 489:89–95. https://doi.org/10.1016/j.bbrc.2017.05.088
Kono M, Hayashi N, Samata T (2000) Molecular mechanism of the nacreous layer formation in Pinctada maxima. Biochem Biophys Res Commun 269:213–218. https://doi.org/10.1006/bbrc.2000.2274
Levi-Kalisman Y, Falini G, Addadi L, Weiner S (2001) Structure of the nacreous organic matrix of a bivalve mollusk shell examined in the hydrated state using cryo-TEM. J Struct Biol 135:8–17. https://doi.org/10.1006/jsbi.2001.4372
Li B, Dewey CN (2011) RSEM: accurate transcript quantification from RNA-Seq data with or without a reference genome. BMC Bioinform 12:323. https://doi.org/10.1186/1471-2105-12-323
Liao Z, Bao L, Fan M et al (2015) In-depth proteomic analysis of nacre, prism, and myostracum of Mytilus shell. J Proteomics 122:26–40. https://doi.org/10.1016/j.jprot.2015.03.027
Liu HL, Liu SF, Ge YJ et al (2007) Identification and characterization of a biomineralization related gene PFMG1 highly expressed in the mantle of Pinctada fucata. Biochemistry 46:844–851. https://doi.org/10.1021/bi061881a
Luo Y-J, Takeuchi T, Koyanagi R et al (2015) The Lingula genome provides insights into brachiopod evolution and the origin of phosphate biomineralization. Nat Commun 6:8301. https://doi.org/10.1038/ncomms9301
Mann K, Poustka AJ, Mann M (2008) The sea urchin (Strongylocentrotus purpuratus) test and spine proteomes. Proteome Sci 6:22. https://doi.org/10.1186/1477-5956-6-22
Marie B, Marie A, Jackson DJ et al (2010) Proteomic analysis of the organic matrix of the abalone Haliotis asinina calcified shell. Proteome Sci 8:54. https://doi.org/10.1186/1477-5956-8-54
Marie B, Trinkler N, Zanella-Cléon I et al (2011) Proteomic identification of novel proteins from the calcifying shell matrix of the Manila clam Venerupis philippinarum. Mar Biotechnol 13:955–962. https://doi.org/10.1007/s10126-010-9357-0 (PMID:21221694)
Marie B, Joubert C, Tayalé A et al (2012) Different secretory repertoires control the biomineralization processes of prism and nacre deposition of the pearl oyster shell. Proc Natl Acad Scie USA 109:20986–20991. https://doi.org/10.1073/pnas.1210552109
Marie B, Jackson DJ, Ramos-Silva P et al (2013) The shell-forming proteome of Lottia gigantea reveals both deep conservations and lineage-specific novelties. FEBS J 280:214–232. https://doi.org/10.1111/febs.12062
Marie B, Arivalagan J, Mathéron L et al (2017) Deep conservation of bivalve nacre proteins highlighted by shell matrix proteomics of the Unionoida Elliptio complanata and Villosa lienosa. J R Soc Interface. https://doi.org/10.1098/rsif.2016.0846
Marin F, Amous R, Guichard N et al (2005) Caspartin and Calprismin, two proteins of the shell calcitic prisms of the Mediterranean fan mussel Pinna nobilis. J Biol Chem 275:20667–20675. https://doi.org/10.1074/jbc.M506526200
Montagnani C, Marie B, Marin F et al (2011) Pmarg-pearlin is a matrix protein involved in nacre framework formation in the pearl oyster Pinctada margaritifera. Chem Bio Chem 12:2033–2043. https://doi.org/10.1002/cbic.201100216
Nagai K, Yano M, Morimoto K, Miyamoto H (2007) Tyrosinase localization in mollusc shells. Comp Biochem Physiol B 146:207–214. https://doi.org/10.1016/j.cbpb.2006.10.105
Naraoka T, Uchisawa H, Mori H et al (2003) Purication, characterization and molecular cloning of tyrosinase from the cephalopod mollusk, Illex argentinus. Eur J Biochem 270:4026–4038. https://doi.org/10.1046/j.1432-1033.2003.03795.x
Nishimura O, Hara Y, Kuraku S (2017) gVolante for standardizing completeness assessment of genome and transcriptome assemblies. Bioinformatics 15(33):3635–3637
Nudelman F, Chen HH, Goldberg HA, Weiner S, Addadi L (2007) Lessons from biomineralization: comparing the growth strategies of mollusk shell prismatic and nacreous layers in Atrina rigida. Faraday Discuss 136:9–25. https://doi.org/10.1039/b704418f
Okumura T, Suzuki M, Nagasawa H, Kogure T (2012) Microstructural variation of biogenic calcite with intracrystalline organic macromolecules. Cryst Growth Des 12:224–230. https://doi.org/10.1021/cg200947c
Petersen TN, Brunak S, von Heijne G, Nielsen H (2011) SignalP 4.0: discriminating signal peptides from transmembrane regions. Nat Methods 8:785–786. https://doi.org/10.1038/nmeth.1701
Ramos-Silva P, Kaandorp J, Huisman L et al (2013) The skeletal proteome of the coral Acropora millepora: the evolution of calcification by co-option and domain shuffling. Mol Biol Evol 30:2099–2112. https://doi.org/10.1093/molbev/mst109
Samata T, Hayashi N, Kono M et al (1999) A new matrix protein family related to the nacreous layer formation of Pinctada fucata. FEBS Lett 462:225–229. https://doi.org/10.1016/s0014-5793(99)01387-3
Samata T, Ikeda D, Kazikawa A et al (2008) A novel phosphorylated glycoprotein in the shell matrix of the oyster Crassostrea nippona. FEBS J 275:2977–2989. https://doi.org/10.1111/j.1742-4658.2008.06453.x
Sánchez-Ferrer A, Rodríguez-López JN, García-Cánovas F, García-Carmona F (1995) Tyrosinase: a comprehensive review of its mechanism. Biochim Biophys Acta 1247(1):1–11. https://doi.org/10.1016/0167-4838(94)00204-t
Sarashina I, Endo K (2001) The complete primary structure of molluscan shell protein 1 (MSP-1), an acidic glycoprotein in the shell matrix of the scallop Patinopecten yessoensis. Mar Biotechnol 3:362–369. https://doi.org/10.1007/s10126-001-0013-6
Schonitzer V, Weiss I (2007) The structure of mollusc larval shells formed in the presence of the chitin synthase inhibitor Nikkomycin Z. BMC Struct Biol 7:71. https://doi.org/10.1186/1472-6807-7-71
Shimizu K, Kimura K, Isowa Y et al (2019) Insights into the evolution of shells and love darts of land snails revealed from their matrix proteins. Genome Biol Evol 11:380–397. https://doi.org/10.1093/gbe/evy242
Simão FA, Waterhouse RM, Ioannidis P, Kriventseva EV, Zdobnov EM (2015) BUSCO: assessing genome assembly and annotation completeness with single-copy orthologs. Bioinformatics 31:3210–3212. https://doi.org/10.1093/bioinformatics/btv351
Stamatakis A (2014) RAxML version 8: A tool for phylogenetic analysis and post-analysis of large phylogenies. Bioinformatics (Oxford, England). https://doi.org/10.1093/bioinformatics/btu033
Suzuki M (2020) Structural and functional analyses of organic molecules regulating biomineralization. Biosci Biotechnol Biochem 84(8):1529–1540. https://doi.org/10.1080/09168451.2020.1762068
Suzuki M, Nagasawa H (2013) Mollusk shell structures and their formation mechanism. Can J Zool 91:349–366. https://doi.org/10.1139/cjz-2012-0333
Suzuki M, Sakuda S, Nagasawa H (2007) Identification of chitin in the prismatic layer of the shell and a chitin synthase gene from the Japanese pearl oyster Pinctada fucata. Biosci Biotechnol Biochem 71(7):1735–1744. https://doi.org/10.1271/bbb.70140
Suzuki M, Saruwatari K, Kogure T et al (2009) An acidic matrix protein, Pif, is a key macromolecule for nacre formation. Science 11:1388–1390. https://doi.org/10.1126/science.1173793
Suzuki M, Iwashima A, Kimura M, Kogure T, Nagasawa H (2013) The molecular evolution of the Pif family proteins in various species of mollusks. Mar Biotechnol 15:145–158. https://doi.org/10.1007/s10126-012-9471-2
Suzuki M, Kogure T, Nagasawa H (2017) Studies on the chemical structures of organic matrices and their functions in the biomineralization processes of molluscan shells. AGri-Biosci Monogr 7:25–39. https://doi.org/10.5047/agbm.2017.00702.0025
Takeuchi T, Sarashina I, Iijima M, Endo K (2008) In vitro regulation of CaCO3 crystal polymorphism by the highly acidic molluscan shell protein Aspein. FEBS Lett 582:591–596. https://doi.org/10.1016/j.febslet.2008.01.026
Takeuchi T, Koyanagi R, Gyoja F et al (2016a) Bivalve-specific gene expansion in the pearl oyster genome: implications of adaptation to a sessile lifestyle. Zool Lett 2:3. https://doi.org/10.1186/s40851-016-0039-2
Takeuchi T, Yamada L, Shinzato C, Sawada H, Satoh N (2016b) Stepwise Evolution of Coral Biomineralization Revealed with Genome-Wide Proteomics and Transcriptomics. PLoS One 11(6):e0156424. https://doi.org/10.1371/journal.pone.0156424
Tamura K, Peterson D, Peterson N et al (2011) MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol Boil Evol 28:2731–2739. https://doi.org/10.1093/molbev/msr121
Thompson JB, Paloczi GT, Kindt JH et al (2000) Direct observation of the transition from calcite to aragonite growth as induced by abalone shell proteins. Biophys J 79:3307–3312. https://doi.org/10.1016/S0006-3495(00)76562-3
Tsukamoto D, Sarashina I, Endo K (2004) Structure and expression of an unusually acidic matrix protein of pearl oyster shells. Biochem Biophys Res Commun 320:1175–1180. https://doi.org/10.1016/j.bbrc.2004.06.072
Yonezawa M, Sakuda S, Yoshimura E, Suzuki M (2016) Molecular cloning and functional analysis of chitinases in the fresh water snail, Lymnaea stagnalis. J Struct Biol 196:107–118. https://doi.org/10.1016/j.jsb.2016.02.021
Zhang C, Xie LP, Huang J, Chen L, Zhang RQ (2006) A novel putative tyrosinase involved in periostracum formation from the pearl oyster (Pinctada fucata). Biochem Biophys Res Commun 342:632–639. https://doi.org/10.1016/j.bbrc.2006.01.182
Zhang G, Fang X, Guo X et al (2012) The oyster genome reveals stress adaptation and complexity of shell formation. Nature 490:49–54. https://doi.org/10.1038/nature11413
Zhao R, Takeuchi T, Luo YJ et al (2018) Dual gene repertoires for larval and adult shells reveal molecules essential for Molluscan shell formation. Mol Biol Evol 35:2751–2761. https://doi.org/10.1093/molbev/msy172
Acknowledgements
We thank Dr Takeshi Takeuchi (Okinawa Institute of Science and Technology Graduate University) for help in annotation of shell matrix proteins and molecular phylogenetic analysis. We also thank Dr Yosuke Ono, Dr Aya Takesono, and Dr Testu Kudoh (University of Exeter) for advice with section in situ hybridization. We also thank Dr Masaei Kanematsu (National Research Institute of Fisheries and Environment of Inland Sea) for the gift of adult specimens of A. pectinata. We thank two professional editors who are both native speakers of English for editing English in this manuscript.
Funding
This research was partially funded by Grant-in-Aid for Scientific Research B (JP19H03045) and Grant-in-Aid for Scientific Research on Innovative Areas IBmS: JSPS KAKENHI Grant Number JP19H05771.
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Shimizu, K., Kintsu, H., Awaji, M. et al. Evolution of Biomineralization Genes in the Prismatic Layer of the Pen Shell Atrina pectinata. J Mol Evol 88, 742–758 (2020). https://doi.org/10.1007/s00239-020-09977-7
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DOI: https://doi.org/10.1007/s00239-020-09977-7