Abstract
Polyploid breeding is widely used in various marine species. Low salinity treatment is an effective method of inducing triploid of bivalve mollusks. In this study, RNA-seq was performed to determine genes and pathways involved in hyposaline adaption and cell division of Pacific oyster (Crassostrea gigas) zygotes, trying to better understand the possible molecular mechanism of hypo-osmotic induction. A total of 26965 unigenes were generated in the de novo assembly of clean Illumina reads with an average length of 934 bp and N50 of 1721 bp. Of 3024 differentially expressed genes (DEGs), 2501 were up-regulated and 523 were down-regulated. GO (Gene Ontology) annotation and KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway analysis of these DEGs revealed that these DEGs participate a variety of biological processes including osmoregulation, cytoskeleton organization, cell survival and death, and substantially modulate cell proliferation and embryonic development. In summery, RNA-seq methodology was applied for the first time to demonstrate hypotonic-induced transcriptomic alteration in oyster zygotes. Our findings not only interpreted the relatively high mortality of induced larvae, but also provided a valuable reference for further investigations on the mechanism of hyposaline induction, thus should aid to the application of low salinity in triploid induction in large scale aquaculture in future.
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Agre, P., 2006. The aquaporin water channels. Proceedings of the American Thoracic Society, 3(1): 5–13, https://doi.org/10.1513/pats.200510-109JH.
Akcha, F., Barranger, A., Bachère, E., Berthelin, C. H., Piquemal, D., Alonso, P., Sallan, R. R., Dimastrogiovanni, G., Porte, C., Menard, D., Szczybelski, A., Benabdelmouna, A., Auffret, M., Rouxel, J., and Burgeot, T., 2016. Effects of an environmentally relevant concentration of diuron on oyster genitors during gametogenesis: Responses of early molecular and cellular markers and physiological impacts. Environmental Science and Pollution Research, 23(8): 8008–8020, https://doi.org/10.1007/s11356-015-5969-2.
Betapudi, V., 2010. Myosin II motor proteins with different functions determine the fate of lamellipodia extension during cell spreading. PLoS One, 5(1): e8560, https://doi.org/10.1371/journal.pone.0008560.
Blerkom, J. V., 1991. Microtubule mediation of cytoplasmic and nuclear maturation during the early stages of resumed meiosis in cultured mouse oocytes. Proceedings of the National Academy of Sciences of the United States of America, 88(11): 5031–5035, https://doi.org/10.1073/pnas.88.11.5031.
Brennan, P., Babbage, J. W., Burgering, B. M. T., Groner, B., Reif, K., and Cantrell, D. A., 1997. Phosphatidylinositol 3-kinase couples the interleukin-2 receptor to the cell cycle regulator E2F. Immunity, 7(5): 679–689, https://doi.org/10.1016/S1074-7613(00)80388-X.
Cao, D., Liu, Y., Ma, L., Jin, X., Guo, G., Tan, R., Liu, Z., Zheng, L., Ye, F., and Liu, W., 2018. Transcriptome analysis of differentially expressed genes involved in selenium accumulation in tea plant (Camellia sinensis). PLoS One, 13(6): e0197 506, https://doi.org/10.1371/journal.pone.0197506.
Chen, K., Li, E., Li, T., Xu, C., Wang, X., Lin, H., Qin, J. G., and Chen, L., 2016. Transcriptome and molecular pathway analysis of the hepatopancreas in the Pacific white shrimp Lito-penaeus vannamei under chronic low-salinity stress. PLoS One, 10(7): e0131503, https://doi.org/10.1371/journal.pone.0131503.
Collart, M. A., and Panasenko, O. O., 2012. The Ccr4-Not complex. Gene, 492(1): 42–53, https://doi.org/10.10162/j.gene.2011.09.033.
Collec, E., Lecomte, M. C., Nemer, W. E., Colin, Y., and Kim, C. L. V., 2011. Novel role for the Lu/BCAM-spectrin interaction in actin cytoskeleton reorganization. Biochemical Journal, 436(3): 699–708, https://doi.org/10.1042/BJ20101717.
Doidge, R., Mittal, S., Aslam, A., and Winkler, G. S., 2012. The anti-proliferative activity of BTG/TOB proteins is mediated via the Caf1a (CNOT7) and Caf1b (CNOT8) deadenylase sub-units of the Ccr4-Not complex. PLoS One, 7(12): e51331, https://doi.org/10.1371/journal.pone.0051331.
Duan, X., and Sun, S., 2018. Actin cytoskeleton dynamics in mammalian oocyte meiosis. Biology of Reproduction, 100(1): 15–24, https://doi.org/10.1093/biolre/ioy163.
Dubrez-Daloz, L., Dupoux, A., and Cartier, J., 2008. IAPS: More than just inhibitors of apoptosis proteins. Cell Cycle, 7(8): 1036–1046, https://doi.org/10.4161/cc.7.8.5783.
Espinoza, J. L., and Minami, M., 2018. Sensing bacterial-induced DNA damaging effects via natural killer group 2 member D immune receptor: From dysbiosis to autoimmunity and carcinogenesis. Frontiers in Immunology, 9: 52, https://doi.org/10.3389/fimmu.2018.00052.
Galluzzi, L., López-Soto, A., Kumar, S., and Kroemer, G., 2016. Caspases connect cell-death signaling to organismal homeostasis. Immunity, 44(2): 221–231, https://doi.org/10.1016/j.immuni.2016.01.020.
Gerard, A., Naciri, Y., Peignon, J. M., Ledu, C., and Phelipot, P., 1994. Optimization of triploid induction by the use of 6-DMAP for the oyster Crassostrea gigas (Thunberg). Aquaculture Research, 25(7): 709–719, https://doi.org/10.1111/j.1365-2109.1994.tb00735.x.
Ghosh, A., Maniloff, J., and Gerling, D. A., 1978. Inhibition of mycoplasma cell division by cytochalasin B. Cell, 13(1): 57–64, https://doi.org/10.1016/0092-8674(78)90137-X.
Guo, Y., Srinivasula, S. M., Druilhe, A., Fernandes-Alnemri, T., and Alnemri, E. S., 2002. Caspase-2 induces apoptosis by releasing proapoptotic proteins from mitochondria. The Journal of Biological Chemistry, 277(16): 13430–13437, https://doi.org/10.1074/jbc.M108029200.
Hamasaki, M., Takeuchi, Y., Miyaki, K., and Yoshizaki, G., 2013. Gonadal development and fertility of triploid grass puffer Ta-kifugu niphobles induced by cold shock treatment. Marine Biotechnology, 15(2): 133–144, https://doi.org/10.1007/s10126-012-9470-3.
Hand, R. E., Nell, J. A., and Thompson, P. A., 2004. Studies on triploid oysters in Australia: XIII. Performance of diploid and triploid Sydney rock oyster, Saccostrea glomerata (Gould, 1850), progeny from a third generation breeding line. Aquaculture, 233(1–4): 93–107, https://doi.org/10.1016/j.aquaculture.2003.09.017.
He, X. Q., Song, Y. Q., Liu, R., Liu, Y., Zhang, F., Zhang, Z., Shen, Y. T., Xu, L., Chen, M. H., Wang, Y. L., Xu, B. H., Yang, X. J., and Wang, H. L., 2016. Axin-1 regulates meiotic spindle organization in mouse oocytes. PLoS One, 11(6): e0157197, https://doi.org/10.1371/journal.pone.0157197.
Hershko, A., and Ciechanover, A., 1998. The ubiquitin system. Annual Review of Biochemistry, 67: 425–479, https://doi.org/10.1146/annurev.biochem.67.1.425.
Houtgraaf, J. H., Versmissen, J., and van der Giessen, W. J., 2006. A concise review of DNA damage checkpoints and repair in mammalian cells. Cardiovascular Revascularization Medicine, 7(3): 165–172, https://doi.org/10.1016/j.carrev.2006.02.002.
Hulata, G., 2001. Genetic manipulations in aquaculture: A review of stock improvement by classical and modern technologies. Genetica, 111(1–3): 155–173, https://doi.org/10.1023/A:1013776931796.
Janku, F., Yap, T. A., and Meric-Bernstam, F., 2018. Targeting the PI3K pathway in cancer: Are we making headway?. Nature Reviews Clinical Oncology, 15(5): 273–291, https://doi.org/10.1038/nrclinonc.2018.28.
Jeung, H. D., Keshavmurthy, S., Lim, H. J., Kim, S. K., and Choi, K. S., 2016. Quantification of reproductive effort of the triploid Pacific oyster, Crassostrea gigas raised in intertidal rack and bag oyster culture system off the west coast of Korea during spawning season. Aquaculture, 464(1): 374–380, https://doi.org/10.1016/j.aquaculture.2016.07.010.
Kanayama, H. O., Tamura, T., Ugai, S., Kagawa, S., Tanahashi, N., Yoshimura, T., Tanaka, K., and Ichihara, A., 1992. Demonstration that a human 26S proteolytic complex consists of a proteasome and multiple associated protein components and hydrolyzes ATP and ubiquitin-ligated proteins by closely linked mechanisms. FEBS Journal, 206(2): 567–578, https://doi.org/10.1111/j.1432-1033.1992.tb16961.x.
Kong, J., Wang, Z. P., Yu, R. H., Liu, J., and Zhang, Y. H., 2011. Triploid induction in Pacific oyster (Crassostrea gigas) by hypotonic treatment and comparison with other induction methods. Journal of Fishery Sciences of China/Zhongguo Shui-chan Kexue, 18(3): 581–587, https://doi.org/10.3724/SP.J.1118.2011.00581.
Liang, L., Deng, L., Chen, Y., Li, G. C., Shao, C., and Tischfield, J. A., 2005. Modulation of DNA end joining by nuclear proteins. The Journal of Biological Chemistry, 280(36): 31442–31449, https://doi.org/10.1074/jbc.M503776200.
Lin, Z., Zhu, X., Zhang, T., You, F., Wu, Z., and Cao, Y., 2016. Effects of hydrostatic pressure on microtubule organization and nucleus changes in gynogenetically activated eggs of olive flounder (Paralichthys olivaceus). Theriogenology, 85(9): 1610–1624, https://doi.org/10.1016/j.theriogenology.2016.01.020.
Livak, K. J., and Schmittgen, T. D., 2001. Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCT method. Methods, 25(4): 402–408, https://doi.org/10.1006/meth.2001.1262.
Lv, J., Liu, P., Wang, Y., Gao, B., Chen, P., and Li, J., 2013. Transcriptome analysis of Portunus trituberculatus in response to salinity stress provides insights into the molecular basis of osmoregulation. PLoS One, 8(12): e82155, https://doi.org/10.1371/journal.pone.0082155.
Lyu, L., Wen, H., Li, Y., Li, J., Zhao, J., Zhang, S., Song, M., and Wang, X., 2018. Deep transcriptomic analysis of black rock-fish (Sebastes schlegelii) provides new insights on responses to acute temperature stress. Scientific Reports, 8(1): 9113, https://doi.org/10.1038/s41598-018-27013-z.
Ma, P., Wang, Z., and Yu, R., 2018. Triploid induction by hyper-osmotic shock in the Yesso scallop, Patinopecten yessoensis. World Aquaculture Society, 50(5): 922–933, https://doi.org/10.1111/jwas.12583.
Meng, H., Terado, T., and Kimura, H., 1998. Apoptosis induced by X-rays and chemical agents in murine fibroblastic cell lines with a defect in repair of DNA double-strand breaks. International Journal of Radiation Biology, 73(5): 503–510, https://doi.org/10.1080/095530098142040.
Meng, J., Zhu, Q., Zhang, L., Li, C., Li, L., She, Z., Huang, B., and Zhang, G., 2013. Genome and transcriptome analyses provide insight into the euryhaline adaptation mechanism of Crassostrea gigas. PLoS One, 8(3): e58563, https://doi.org/10.1371/journal.pone.0058563.
Mininni, A. N., Milan, M., Ferraresso, S., Petochi, T., Marco, P. D., Marino, G., Livi, S., Romualdi, C., Bargelloni, L., and Patarnello, T., 2014. Liver transcriptome analysis in gilthead sea bream upon exposure to low temperature. BMC Genomics, 15: 765, https://doi.org/10.1186/1471-2164-15-765.
Nell, J. A., Hand, R. E., Goard, L. J., McAdam, S. P., and Maguire, G. B., 1996. Studies on triploid oysters in Australia: Evaluation of cytochalasin B and 6-dimethylaminopurine for triploidy induction in Sydney rock oysters Saccostrea com-mercialis (Iredale and Roughley). Aquaculture Research, 27(9): 689–698, https://doi.org/10.1046/j.1365-2109.1996.t01-1-00777.x.
Normand, J., Ernande, B., Haure, J., McCombie, H., and Boudry, P., 2009. Reproductive effort and growth in Crassostrea gigas: Comparison of young diploid and triploid oysters issued from natural crosses or chemical induction. Aquatic Biology, 7(3): 229–241, https://doi.org/10.3354/ab00190.
Okumura, S. I., Arai, K., Harigaya, Y., Eguchi, H., Sakai, M., Senbokuya, H., Furukawa, S., and Yamamori, K., 2007. Highly efficient induction of triploid Pacific abalone Haliotis discus hannai by caffeine treatment. Fisheries Science, 73(2): 237–243, https://doi.org/10.1111/j.1444-2906.2007.01329.x.
Pierce, S. K., and Amende, L. M., 1981. Control mechanisms of amino acid-mediated cell volume regulation in salinity-stressed molluscs. Journal of Experimental Zoology, 215(3): 247–257, https://doi.org/10.1002/jez.1402150304.
Piferrer, F., Beaumont, A., Falguière, J. C., Flajshans, M., Haffray, P., and Colombo, L., 2009. Polyploid fish and shellfish: Production, biology and applications to aquaculture for performance improvement and genetic containment. Aquaculture, 293(3–4): 125–156, https://doi.org/10.1016/j.aquaculture.2009.04.036.
Reardon, J. T., and Sancar, A., 2006. Purification and characterization of Escherichia coli and human nucleotide excision repair enzyme systems. Methods in Enzymology, 408: 189–213, https://doi.org/10.1016/S0076-6879(06)08012-8.
Rime, H., Neant, I., Guerrier, P., and Ozon, R., 1989. 6-Dime-thylaminopurine (6-DMAP), a reversible inhibitor of the transition to metaphase during the first meiotic cell division of the mouse oocyte. Developmental Biology, 133(1): 169–179, https://doi.org/10.1016/0012-1606(89)90308-4.
Sangiao-Alvarellos, S., Laiz-Carrión, R., Guzmán, J. M., del Río, M. P. M., Miguez, J. M., Mancera, J. M., and Soengas, J. L., 2003. Acclimation of S. aurata to various salinities alters energy metabolism of osmoregulatory and nonosmoregulatory organs. American Journal of Physiology-Regulatory, Integrative and Comparative Physiology, 285(4): R897–R907, https://doi.org/10.1152/ajpregu.00161.2003.
Sarà, G., Reid, G. K., Rinaldi, A., Palmeri, V., Troell, M., and Kooijman, S. A. L. M., 2012. Growth and reproductive simulation of candidate shellfish species at fish cages in the Southern Mediterranean: Dynamic Energy Budget (DEB) modelling for integrated multi-trophic aquaculture. Aquaculture, 324–325(12): 259–266, https://doi.org/10.1016/j.aquaculture.2011.10.042.
Schauber, C., Chen, L., Tongaonkar, P., Vega, I., Lambertson, D., Potts, W., and Madura, K., 1998. Rad23 links DNA repair to the ubiquitin/proteasome pathway. Nature, 391(6668): 715–718, https://doi.org/10.1038/35661.
Si, Y., Wen, H., Li, Y., He, F., Li, J., Li, S., and He, H., 2018. Liver transcriptome analysis reveals extensive transcriptional plasticity during acclimation to low salinity in Cynoglossus semilaevis. BMC Genomics, 19(1): 464, https://doi.org/10.1186/s12864-018-4825-4.
Tournaire-Roux, C., Sutka, M., Javot, H., Gout, E., Gerbeau, P., Luu, D. T., Bligny, R., and Maurel, C., 2003. Cytosolic pH regulates root water transport during anoxic stress through gating of aquaporins. Nature, 425(6956): 393–397, https://doi.org/10.1038/nature01853.
Umenishi, F., and Schrier, R. W., 2003. Hypertonicity-induced aquaporin-1 (AQP1) expression is mediated by the activation of MAPK pathways and hypertonicity responsive element in the AQP1 gene. The Journal of Biological Chemistry, 278(18): 15765–15770, https://doi.org/10.1074/jbc.M2099 80200.
Vong, Q. P., Cao, K., Li, H. Y., Iglesias, P. A., and Zheng, Y., 2005. Chromosome alignment and segregation regulated by ubiquitination of survivin. Science, 310(5753): 1499–1504, https://doi.org/10.1126/science.1120160.
Wang, W., Teng, F., Lin, Y., Ji, D., Xu, Y., Chen, C., and Xie, C., 2018. Transcriptomic study to understand thermal adaptation in a high temperature-tolerant strain of Pyropia haitanensis. PLoS One, 13(4): e0195842, https://doi.org/10.1371/journal.pone.0195842.
Wang, Y., Guo, X., DeBrosse, G., and Ford, S., 2005. Superior growth in natural triploid eastern oyster produced by diploid x tetraploid crosses. Journal of Shellfish Research, 24(4): 1274.
Williams, M. R., Arthur, J. S. C., Balendran, A., van der Kaay, J., Poli, V., Cohen, P., and Alessi, D. R., 2000. The role of 3-phosphoinositide-dependent protein kinase 1 in activating AGC kinases defined in embryonic stem cells. Current Biology, 10(8): 439–448, https://doi.org/10.1016/S0960-9822(00)00441-3.
Windisch, H. S., Frickenhaus, S., John, U., Knust, R., Pörtner, H. O., and Lucassen, M., 2014. Stress response or beneficial temperature acclimation: Transcriptomic signatures in Antarctic fish (Pachycara brachycephalum). Molecular Ecology, 23(14): 3469–3482, https://doi.org/10.1111/mec.12822.
Xu, Z., Gan, L., Li, T., Xu, C., Chen, K., Wang, X., Qin, J. G., Chen, L., and Li, E., 2015. Transcriptome profiling and molecular pathway analysis of genes in association with salinity adaptation in Nile Tilapia Oreochromis niloticus. PLoS One, 10(8): e0136506, https://doi.org/10.1371/journal.pone.0136506.
Yan, L., Li, Y., Wang, Z., Su, J., Yu, R., Yan, X., Ma, P., and Cui, Y., 2018. Stress response to low temperature: Transcriptomic characterization in Crassostrea sikamea × Crassostrea angu-lata hybrids. Aquaculture Research, 49(10): 3374–3385, https://doi.org/10.1111/are.13801.
Yan, L., Su, J., Wang, Z., Yan, X., Yu, R., Ma, P., Li, Y., and Du, J., 2017. Transcriptomic analysis of Crassostrea sikamea × Crassostrea angulata hybrids in response to low salinity stress. PLoS One, 12(2): e0171483, https://doi.org/10.1371/journal.pone.0171483.
Yuan, Y., Chen, J., Chen, B., Qu, K., Guo, F., Li, Q., and Cui, Y., 2000. Study on adaptability of scallop Chlamys farreri to environment-effects of salinity and pH on survival, respiration, ingestion and digestion. Journal of Fishery Sciences of China, 7(4): 73–77.
Zeng, J., Kamiyama, T., Niwa, R., and King-Jones, K., 2018. The Drosophila CCR4-NOT complex is required for cholesterol homeostasis and steroid hormone synthesis. Developmental Biology, 443(1): 10–18, https://doi.org/10.1016/j.ydbio.2018.08.012.
Zhang, D., Wang, F., Dong, S., and Lu, Y., 2016. De novo assembly and transcriptome analysis of osmoregulation in Lito-penaeus vannamei under three cultivated conditions with different salinities. Gene, 578(2): 185–193, https://doi.org/10.1016/j.gene.2015.12.026.
Zhang, F., Zhang, Z., Kong, D., Zhang, X., Chen, L., Zhu, X., Lu, Y., and Zheng, S., 2014. Tetramethylpyrazine reduces glucose and insulin-induced activation of hepatic stellate cells by inhibiting insulin receptor-mediated PI3K/AKT and ERK pathways. Molecular and Cellular Endocrinology, 382(1): 197–204, https://doi.org/10.1016/j.mce.2013.09.020.
Zhang, G., Chang, Y., Song, J., Ding, J., and Shen, J, 1998. Tri-ploidy induction in Pacific oyster Crassostrea gigas by caffeine with thermal shock. Chinese Journal of Oceanology and Limnology, 16(3): 249–255, https://doi.org/10.1007/BF02848731.
Zhang, L., Feng, Q., Sun, L., Ding, K., Huo, D., Fang, Y., Zhang, T., and Yang, H., 2018. Differential gene expression in the intestine of sea cucumber (Apostichopus japonicus) under low and high salinity conditions. Comparative Biochemistry and Physiology — Part D, 25: 34–41, https://doi.org/10.1016/j.cbd.2017.11.001.
Zhang, L., Yang, X., Qi, X., Guo, C., and Jing, Z., 2017. Characterizing the transcriptome and microsatellite markers for almond (Amygdalus communis L.) using the Illumina sequencing platform. Hereditas, 155(1): 14, https://doi.org/10.1186/s41065-017-0049-x.
Zhang, X., Li, F., and Xiang, J., 2003. Chromosome behavior of heat shock induced triploid in Fenneropenaeus chinensis. Chinese Journal of Oceanology and Limnology, 21(3): 222–228, https://doi.org/10.1007/BF02842838.
Zhang, X., Wen, H., Wang, H., Ren, Y., Zhao, J., and Li, Y., 2017. RNA-seq analysis of salinity stress-responsive transcriptome in the liver of spotted sea bass (Lateolabrax maculatus). PLoS One, 12(3): e0173238, https://doi.org/10.1371/journal.pone.0173238.
Zhao, Q., Pan, L., Ren, Q., and Hu, D., 2015. Digital gene expression analysis in hemocytes of the white shrimp Litope-naeus vannamei in response to low salinity stress. Fish & Shellfish Immunology, 42(2): 400–407, https://doi.org/10.1016/j.fsi.2014.11.020.
Zhao, X., Yu, H., Kong, L., and Li, Q., 2012. Transcriptomic responses to salinity stress in the Pacific Oyster Crassostrea gigas. PLoS One, 7(9): e46244, https://doi.org/10.1371/journal.pone.0046244.
Zurburg, W., and Zwaan, A. D., 1981. The role of amino acids in anaerobiosis and osmoregulation in bivalves. Journal of Experimental Zoology, 215(3): 315–325, https://doi.org/10.1002/jez.1402150309.
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This work was supported by the National Natural Science Foundation of China (No. 31172403). We are grateful to Drs. Tingting Wang and Zongwu Zhao for their helpful suggestions and writing assistance.
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Li, Y., Wang, Z., Cui, Y. et al. Transcriptomic Analysis of Pacific Oyster (Crassostrea gigas) Zygotes Under Hypotonic Triploid Induction. J. Ocean Univ. China 20, 147–158 (2021). https://doi.org/10.1007/s11802-021-4450-8
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DOI: https://doi.org/10.1007/s11802-021-4450-8