Cite this paper:
WU Xiangwei, LIU Xiande, YU Ziniu. Analysis of novel immune-related genes and microsatellite markers in the transcriptome of Paphia undulata[J]. HaiyangYuHuZhao, 2019, 37(4): 1301-1316

Analysis of novel immune-related genes and microsatellite markers in the transcriptome of Paphia undulata

WU Xiangwei1,2,4, LIU Xiande3, YU Ziniu1,4
1 Key Laboratory of Tropical Marine Bio-resources and Ecology, Guangdong Provincial Key Laboratory of Applied Marine Biology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, uangzhou 510301, China;
2 Animal Science and Technology College, Yunnan Agricultural University, Kunming 650201, China;
3 Key Laboratory of Healthy Mariculture for the East China Sea, Ministry of Agriculture of China; Fisheries College, Jimei University, Xiamen 361021, China;
4 University of Chinese Academy of Sciences, Beijing 100049, China
Abstract:
Increasingly, exogenous stressors such as pathogen infections, variable water conditions, and pollution are resulting in high mortality of Paphia undulata, deleteriously affecting the quality of clam harvests. The foot is a burrowing organ in clams. Physical damage and constant contact with the external environment cause the foot to be highly sensitive to pathogen invasion and water condition variation. In the present study, the foot tissue transcriptome was analyzed to identify genes involved in immune and stress responses. The P. undulata transcriptome included 5 286 668 078 bp reads generated by Illumina Hiseq 2000 sequencing and were assembled into 1 785 226 contigs by de novo method. The contigs were clustered into 99 339 transcripts and further grouped into 60 201 unigenes. Of them, 22 260 unigenes were successfully annotated using public databases. Twelve genes that were response to immune and stress were identified with abundant expression levels, including heat shock protein 70, cold shock protein, complement C3, cathepsin L, ubiquitin carboxyl-terminal hydrolase L5, and translationally controlled tumor protein. Furthermore, 566 unigenes were found homologous to genes involved in the immune response systems of pathogen discrimination, signal transduction, and immune effector, such as lectins, toll-like receptors, complement pathway, toll-like receptor signaling pathway, heat shock proteins, antioxidant enzymes, lysozymes, and mucins, indicating that P. undulata could have a complete set of innate immune mechanisms. In addition, 4 270 microsatellite markers (SSRs) were identified from 60 201 unigenes, of which trinucleotide repeats were most abundant and 16 SSRs were tested to be polymorphic. The present study provides a new insight into innate immunity and stress response mechanisms in P. undulata.
Key words:    Paphia undulata|foot tissue|transcriptome|innate immunity|unigene|microsatellite   
Received: 2018-05-21   Revised: 2018-07-11
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References:
Aguirre J, Ríos-Momberg M, Hewitt D, Hansberg, W. 2005.Reactive oxygen species and development in microbial eukaryotes. Trends Microbiol., 13(3):111-118.
Akira S. 2003. Toll-like receptor signaling. J. Biol. Chem., 278(40):38 105-38 108.
Altschul S F, Madden T L, Schäffer A, Zhang J H, Zhang Z, Miller W, Lipman D J. 1997. Gapped BLAST and PSIBLAST:a new generation of protein database search programs. Nucleic Acids Res., 25(17):3 389-3 402.
An H S, Lee J W. 2012. Development of microsatellite markers for the Korean mussel, Mytilus coruscus (Mytilidae)using next-generation sequencing. Int. J. Mol. Sci., 13(8):10 583-10 593.
Bachali S, Jager M, Hassanin A, Schoentgen F, Jollès P, FialaMedioni A, Deutsch J S. 2002. Phylogenetic analysis of invertebrate lysozymes and the evolution of lysozyme function. J. Mol. Evol., 54(5):652-664.
Bahia A C, Kubota M S, Tempone A J, Pinheiro W D, Tadei W P, Secundino N F C, Traub-Csekö Y M, Pimenta P F P. 2010. Anopheles aquasalis infected by Plasmodium vivax displays unique gene expression profiles when compared to other malaria vectors and plasmodia. PLoS One, 5(3):e9795.
Beaz-Hidalgo R, Balboa S, Romalde J L, Figueras M J. 2010.Diversity and pathogenecity of Vibrio species in cultured bivalve molluscs. Environ. Microbiol. Rep., 2(1):34-43.
Bettencourt R, Pinheiro M, Egas C, Gomes P, Afonso M, Shank T, Santos R S. 2010. High-throughput sequencing and analysis of the gill tissue transcriptome from the deep-sea hydrothermal vent mussel Bathymodiolus azoricus. BMC Genomics, 11:559.
Botstein D, White R L, Skolnick M, Davis R W. 1980.Construction of a genetic linkage map in man using restriction fragment length polymorphisms. Am. J. Hum.Genet., 32(3):314-331.
Brun N T, Bricelj V M, MacRae T H, Ross N W. 2008. Heat shock protein responses in thermally stressed bay scallops, Argopecten irradians, and sea scallops, Placopecten magellanicus. J. Exp. Mar. Biol. Ecol., 358(2):151-162.
Carroll M C. 2004. The complement system in regulation of adaptive immunity. Nat. Immunol., 5(10):981-986.
Chen X M, Li J K, Xiao S J, Liu X D. 2016. De novo assembly and characterization of foot transcriptome and microsatellite marker development for Paphia textile.Gene, 576(1):537-543.
Clark M S, Thorne M A S, Vieira F A, Cardoso J C R, Power D M, Peck L S. 2010. Insights into shell deposition in the Antarctic bivalve Laternula elliptica:gene discovery in the mantle transcriptome using 454 pyrosequencing. BMC Genomics, 11:362.
Conesa A, Götz S, García-Gómez J M, Terol J, Talón M, Robles M. 2005. Blast2GO:a universal tool for annotation, visualization and analysis in functional genomics research. Bioinformatics, 21(18):3 674-3 676.
Cronin S J, Nehme N T, Limmer S, Liegeois S, Pospisilik J A, Schramek D, Leibbrandt A, Simoes R D M, Gruber S, Puc U, Ebersberger I, Zoranovic T, Neely G G, von Haeseler A, Ferrandon D, Penninger J M. 2009. Genome-wide RNAi screen identifies genes involved in intestinal pathogenic bacterial infection. Science, 325(5938):340-343.
Delanghe J R, Speeckaert R, Speeckaert M M. 2014.Complement C3 and its polymorphism:biological and clinical consequences. Pathology, 46(1):1-10.
Deng Y W, Lei Q N, Tian Q L, Xie S H, Du X D, Li J H, Wang L Q, Xiang Y X. 2014. De novo assembly, gene annotation, and simple sequence repeat marker development using Illumina paired-end transcriptome sequences in the pearl oyster Pinctada maxima. Biosci. Biotechnol. Biochem., 78(10):1 685-1 692.
Dunkelberger J R, Song W C. 2010. Complement and its role in innate and adaptive immune responses. Cell Res., 20(1):34-50.
Feldmeyer B, Wheat C W, Krezdorn N, Rotter B, Pfenninger M. 2011. Short read Illumina data for the de novo assembly of a non-model snail species transcriptome(Radix balthica, Basommatophora, Pulmonata), and a comparison of assembler performance. BMC Genomics, 12:317.
Feng B B, Dong L L, Niu D H, Meng S S, Zhang B, Liu D B, Hu S N, Li J L. 2010. Identification of immune genes of the Agamaki clam (Sinonovacula constricta) by sequencing and bioinformatic analysis of ESTs. Mar.Biotechnol., 12(3):282-291.
Gendler S J, Spicer A P. 1995. Epithelial mucin genes. Annu.Rev. Physiol., 57:607-634.
Gerdol M, Manfrin C, De Moro G, Figueras A, Novoa B, Venier P, Pallavicini A. 2011. The C1q domain containing proteins of the Mediterranean mussel Mytilus galloprovincialis:a widespread and diverse family of immune-related molecules. Dev. Comp. Immunol., 35(6):635-643.
Gourdine J P, Smith-Ravin E J. 2007. Analysis of a cDNAderived sequence of a novel mannose-binding lectin, codakine, from the tropical clam Codakia orbicularis.Fish Shellfish Immunol., 22(5):498-509.
Grieshaber M K, Hardewig I, Kreutzer U, Pörlner H O. 1994.Physiological and metabolic responses to hypoxia in invertebrates. In:Reviews of Physiology, Biochemistry and Pharmacology. Springer, Berlin, Heidelberg. p.43-147.
Gueguen Y, Cadoret J P, Flament D, Barreau-Roumiguière C, Girardot A L, Garnier J, Hoareau A, Bachère E, Escoubas J M. 2003. Immune gene discovery by expressed sequence tags generated from hemocytes of the bacteria-challenged oyster, Crassostrea gigas. Gene, 303:139-145.
Haas B J, Papanicolaou A, Yassour M, Grabherr M, Blood P D, Bowden J, Couger M B, Eccles D, Li B, Lieber M, MacManes M D, Ott M, Orvis J, Pochet N, Strozzi F, Weeks N, Westerman R, William T, Dewey C N, Henschel R, LeDuc R D, Friedman N, Regev A. 2013. De novo transcript sequence reconstruction from RNA-seq using the Trinity platform for reference generation and analysis.Nat. Protoc., 8(8):1 494-1 512.
Huan P, Wang H X, Liu B Z. 2012. Transcriptomic analysis of the clam Meretrix meretrix on different larval stages. Mar.Biotechnol., 14(1):69-78.
Iwanaga S, Lee B L. 2005. Recent advances in the innate immunity of invertebrate animals. J. Biochem. Mol. Boil., 38(2):128-150.
Iwasaki A, Medzhitov R. 2004. Toll-like receptor control of the adaptive immune responses. Nat. Immunol., 5(10):987-995.
Janeway C A Jr, Medzhitov R. 2002. Innate immune recognition. Annu. Rev. Immunol., 20:197-216.
Jensen K T, Castro N F, Bachelet G. 1999. Infectivity of Himasthla spp. (Trematoda) in cockle (Cerastoderma edule) spat. J. Mar. Biol. Assoc. UK, 79(2):265-271.
Jeong E, Lee J Y. 2011. Intrinsic and extrinsic regulation of innate immune receptors. Yonsei Med. J., 52(3):379-392.
Jiang WN, Hon Y, Inouye M. 1997. CspA, the major coldshock protein of Escherichia coli, is an RNA chaperone.J. Biol. Chem., 272(1):196-202.
Kim J Y, Adhya M, Cho S K, Choi K S, Cho M. 2008.Characterization, tissue expression, and immunohistochemical localization of MCL3, a C-type lectin produced by Perkinsus olseni-infected Manila clams (Ruditapes philippinarum). Fish Shellfish Immunol., 25(5):598-603.
Lauckner G. 1983. Diseases of Mollusca:Bivalvia. In:Kinne O ed. Diseases of Marine Animals. Biologische Anstalt Helgoland, Hamburg, Germany. p.477-961.
Leethochavalit S, Chalermwat K, Upatham E S, Choi K S, Sawangwong P, Kruatrachue M. 2004. Occurrence of Perkinsus sp. in undulated surf clams Paphia undulata from the Gulf of Thailand. Dis. Aquat. Org., 60(2):165-171.
Li H J, Liu W D, Gao X G, Zhu D, Wang J, Li Y F, He C B. 2011. Identification of host-defense genes and development of microsatellite markers from ESTs of hard clam Meretrix meretrix. Mol. Biol. Rep., 38(2):769-775.
Loker E S, Adema C M, Zhang S M, Kepler T B. 2004.Invertebrate immune systems- not homogeneous, not simple, not well understood. Immunol. Rev., 198(1):10-24.
Matsubara H, Ogawa T, Muramoto K. 2006. Structures and functions of C-type lectins in marine invertebrates.Tohoku J. Agric. Res., 57(1-2):71-86.
Moreira R, Balseiro P, Planas J V, Fuste B, Beltran S, Novoa B, Figueras A. 2012. Transcriptomics of in vitro immunestimulated hemocytes from the Manila clam Ruditapes philippinarum using high-throughput sequencing. PLoS One, 7(4):e35009.
Mouritsen K N, Poulin R. 2003. The risk of being at the top:foot-cropping in the New Zealand cockle Austrovenus stutchburyi. J. Mar. Biol. Assoc. UK, 83(3):497-498.
Niu D H, Jin K, Wang L, Feng B B, Li J L. 2013b. Molecular characterization and expression analysis of four cathepsin L genes in the razor clam, Sinonovacula constricta. Fish Shellfish Immunol., 35(2):581-588.
Niu D H, Wang L, Sun F Y, Liu Z J, Li J L. 2013a. Development of molecular resources for an intertidal clam, Sinonovacula constricta, using 454 transcriptome sequencing. PLoS One, 8(7):e67456.
O'Connell-Milne S, Poulin R, Savage C, Rayment W. 2016.Reduced growth, body condition and foot length of the bivalve Austrovenus stutchburyi in response to parasite infection. J. Exp. Mar. Biol. Ecol., 474:23-28.
Pallavicini A, del Mar Costa M, Gestal C, Dreos R, Figueras A, Venier P, Novoa B. 2008. High sequence variability of myticin transcripts in hemocytes of immune-stimulated mussels suggests ancient host-pathogen interactions. Dev.Comp. Immunol., 32(3):213-226.
Parcellier A, Gurbuxani S, Schmitt E, Solary E, Garrido C. 2003. Heat shock proteins, cellular chaperones that modulate mitochondrial cell death pathways. Biochem.Biophys. Res. Commun., 304(3):505-512.
Paul-Pont I, de Montaudouin X, Gonzalez P, Soudant P, Baudrimont M. 2010. How life history contributes to stress response in the Manila clam Ruditapes philippinarum. Environ. Sci. Pollut. Res., 17(4):987-998.
Peng M X, Niu D H, Wang F, Chen Z Y, Li J L. 2016.Complement C3 gene:Expression characterization and innate immune response in razor clam Sinonovacula constricta. Fish Shellfish Immunol., 55:223-232.
Philipp E, Pörtner HO, Abele D. 2005. Mitochondrial ageing of a polar and a temperate mud clam. Mech. Ageing Dev., 126(5):610-619.
Ruddell R G, Hoang-Le D, Barwood J M, Rutherford P S, Piva T J, Watters D J, Santambrogio P, Arosio P, Ramm G A. 2009. Ferritin functions as a proinflammatory cytokine via iron-independent protein kinase C zeta/nuclear factor kappaB-regulated signaling in rat hepatic stellate cells.Hepatology, 49(3):887-900.
Sambrook J, Fritsch E F, Maniatis J. 1989. Molecular Cloning:A Laboratory Manual. 2nd edn. Cold Spring Harbor Laboratory Press, New York. 1625p.
Shi M J, Lin Y, Xu G R, Xie L P, Hu X L, Bao Z M, Zhang R Q. 2013. Characterization of the Zhikong scallop(Chlamys farreri) mantle transcriptome and identification of biomineralization-related genes. Mar. Biotechnol., 15(6):706-715.
Song L S, Wang L L, Qiu L M, Zhang H A. 2011. Bivalve immunity. In:Söderhäll K ed. Invertebrate Immunity.Springer, Boston, MA. p.44-65.
Song X Y, Zhang H, Zhao J M, Wang L L, Qiu L M, Mu C K, Liu X L, Qiu L H, Song L S. 2010. An immune responsive multidomain galectin from bay scallop Argopectens irradians. Fish Shellfish Immunol., 28(2):326-332.
Takeda K, Akira S. 2005. Toll-like receptors in innate immunity. Int. Immunol., 17(1):1-14.
Tatusov R L, Galperin M Y, Natale D A, Koonin E V. 2000.The COG database:a tool for genome-scale analysis of protein functions and evolution. Nucleic Acids Res., 28(1):33-36.
Thieringer H A, Jones P G, Inouye M. 1998. Cold shock and adaptation. Bioessays, 20(1):49-57.
Thomas F, Renaud F, de Meeûs T, Poulin R. 1998. Manipulation of host behaviour by parasites:ecosystem engineering in the intertidal zone? Proc. Roy. Soc. B:Biol. Sci., 265(1401):1 091-1 096.
Underhill D M, Ozinsky A. 2002. Toll-like receptors:key mediators of microbe detection. Curr. Opin. Immunol., 14(1):103-110.
van de Wetering J K, van Golde L M G, Batenburg J J. 2004.Collectins:players of the innate immune system. FEBS J., 271(7):1 229-1 249.
Venier P, Varotto L, Rosani U, Millino C, Celegato B, Bernante F, Lanfranchi G, Novoa B, Roch P, Figueras A, Pallavicini A. 2011. Insights into the innate immunity of the Mediterranean mussel Mytilus galloprovincialis. BMC Genomics, 12:69.
Wang A M, Wang Y, Gu Z F, Li S F, Shi Y H, Guo X M. 2011.Development of expressed Sequence tags from the pearl oyster, Pinctada martensii Dunker. Mar. Biotechnol., 13(2):275-283.
Wang L L, Qiu L M, Zhou Z, Song L S. 2013. Research progress on the mollusc immunity in China. Dev. Comp.Immunol., 39(1-2):2-10.
Wang L L, Song L S, Zhao J M, Qiu L M, Zhang H, Xu W, Li H L, Li C H, Wu L T, Guo X M. 2009. Expressed sequence tags from the Zhikong scallop (Chlamys farreri):discovery and annotation of hostdefense genes. Fish Shellfish Immunol., 26(5):744-750.
Weber M H W, Fricke I, Doll N, Marahiel M A. 2002.CSDBase:an interactive database for cold shock domaincontaining proteins and the bacterial cold shock response.Nucleic Acids Res., 30(1):375-378.
Wu X W, Tan J, Cai M Y, Liu X D. 2014. Molecular cloning, characterization, and expression analysis of a heat shock protein (HSP) 70 gene from Paphia undulata. Gene, 543(2):275-285.
Yang C Y, Wang L L, Siva V S, Shi X W, Jiang Q F, Wang J J, Zhang H, Song L S. 2012. A novel cold-regulated cold shock domain containing protein from scallop Chlamys farreri with nucleic acid-binding activity. PLoS One, 7(2):e32012.
Yeh F C, Yang R, Boyle T J, Ye Z, Xiyan J M. 2000. PopGene32, Microsoft Windows-based freeware for population genetic analysis, version 1.32. Molecular Biology and Biotechnology Centre, University of Alberta, Edmonton, Alberta, Canada.
Yeh H Y, Klesius P H. 2010. Characterization and tissue expression of channel catfish (Ictalurus punctatus Rafinesque, 1818) ubiquitin carboxyl-terminal hydrolase L5 (UCHL5) cDNA. Mol. Biol. Rep., 37(3):1 229-1 234.
Zhang H, Song L S, Li C H, Zhao J M, Wang H, Qiu L M, Ni D J, Zhang Y. 2008. A novel C1q-domain-containing protein from Zhikong scallop Chlamys farreri with lipopolysaccharide binding activity. Fish Shellfish Immunol., 25(3):281-289.