Molecular identification, expression pattern, and in-vitro bioactivity analysis of insulin-like growth factor 2 in olive flounder <i>Paralichthys olivaceus</i> -- Journal of Oceanology and Limnology,2023,41(1):280-289

	Cite this paper:
	Shuang JIAO, Xungang TAN, Feng YOU, Shujing ZHANG, Qiuxiang PANG. Molecular identification, expression pattern, and in-vitro bioactivity analysis of insulin-like growth factor 2 in olive flounder Paralichthys olivaceus[J]. Journal of Oceanology and Limnology, 2023, 41(1): 280-289

Molecular identification, expression pattern, and in-vitro bioactivity analysis of insulin-like growth factor 2 in olive flounder Paralichthys olivaceus

Shuang JIAO¹, Xungang TAN^2,3, Feng YOU^2,3, Shujing ZHANG¹, Qiuxiang PANG¹

1 Anti-Aging&Regenerative Medicine Research Institution, School of Life Sciences and Medicine, Shandong University of Technology, Zibo 255000, China;
2 CAS and Shandong Province Key Laboratory of Experimental Marine Biology, Center for Ocean Mega-Science, Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, China;
3 Laboratory for Marine Biology and Biotechnology, Pilot National Laboratory for Marine Science and Technology(Qingdao), Qingdao 266237, China

Abstract:

Insulin-like growth factors (IGFs) are key regulators of development and growth. Here, we characterized the igf2 gene from olive flounder (Paralichthys olivaceus) and determined its temporal and spatial expression. We set up an in-vitro protein expression system in eukaryotic human embryonic kidney (HEK293T) cells and explored its effects on cell proliferation. The flounder igf2 cDNA contained a 648-bp open reading frame (ORF) encoding a protein of 215 amino acids (aa), which spanned the complete signal peptide (47 aa), mature peptide (70 aa), and E domain (98 aa). In adult flounder, igf2 mRNA was detected in all selected tissues. In early development, igf2 mRNA was detected throughout development from unfertilized eggs to hatching-stage embryos. In-situ hybridization analysis indicated that igf2 mRNA was specially expressed in the brain region, floor plate, hypochord, otic vesicle, and pectoral fin during embryogenesis. Western blotting analysis indicated that the soluble recombinant flounder IGF2 protein was successfully produced through eukaryotic expression in HEK293T cells. In addition, the recombinant IGF2 protein significantly promoted the proliferation of human cervical carcinoma (HeLa) and HEK293T cells. These results provide new information about the structural and functional conservation, expression patterns, and biological activity of the igf2 in teleosts.

Key words: aquaculture|flatfish|igf2|proliferation

Received: 2021-10-29 Revised:

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References:
Ayson F G, De Jesus E G, Moriyama S et al. 2002. Differential expression of insulin-like growth factor I and II mRNAs during embryogenesis and early larval development in rabbitfish, Siganus guttatus. General and Comparative Endocrinology, 126(2): 165-174, https://doi.org/10.1006/gcen.2002.7788.
Bäck K, Brännmark C, Strålfors P et al. 2011. Differential effects of IGF-I, IGF-II and insulin in human preadipocytes and adipocytes--role of insulin and IGF-I receptors. Molecular and Cellular Endocrinology, 339(1-2): 130-135, https://doi.org/10.1016/j.mce.2011.04.005.
Bendall S C, Stewart M H, Menendez P et al. 2007. IGF and FGF cooperatively establish the regulatory stem cell niche of pluripotent human cells in vitro. Nature, 448(7157):1015-1021, https://doi.org/10.1038/nature06027.
Brown A L, Graham D E, Nissley S P et al. 1986. Developmental regulation of insulin-like growth factor II mRNA in different rat tissues. Journal of Biological Chemistry, 261(28): 13144-13150.
Caelers A, Berishvili G, Meli M L et al. 2004. Establishment of a real-time RT-PCR for the determination of absolute amounts of IGF-I and IGF-II gene expression in liver and extrahepatic sites of the tilapia. General and Comparative Endocrinology, 137(2): 196-204, https://doi.org/10.1016/j.ygcen.2004.03.006.
Chen J Y, Chang C Y, Chen J C et al. 1997. Production of biologically active recombinant tilapia insulin-like growth factor-II polypeptides in Escherichia coli cells and characterization of the genomic structure of the coding region. DNA and Cell Biology, 16(7): 883-892, https://doi.org/10.1089/dna.1997.16.883.
Codina M, De La Serrana D G, Sánchez-Gurmaches J et al. 2008.Metabolic and mitogenic effects of IGF-II in rainbow trout(Oncorhynchus mykiss) myocytes in culture and the role of IGF-II in the PI3K/Akt and MAPK signalling pathways.General and Comparative Endocrinology, 157(2): 116-124, https://doi.org/10.1016/j.ygcen.2008.04.009.
Denley A, Cosgrove L J, Booker G W et al. 2005. Molecular interactions of the IGF system. Cytokine & Growth Factor Reviews, 16(4-5): 421-439, https://doi.org/10.1016/j.cytogfr.2005.04.004.
Duan C M, Ding J, Li Q et al. 1999. Insulin-like growth factor binding protein 2 is a growth inhibitory protein conserved in zebrafish. Proceedings of the National Academy of Sciences of the United States of America, 96(26): 15274-15279, https://doi.org/10.1073/pnas.96.26.15274.
Duan C M, Ren H X, Gao S. 2010. Insulin-like growth factors(IGFs), IGF receptors, and IGF-binding proteins: roles in skeletal muscle growth and differentiation. General and Comparative Endocrinology, 167(3): 344-351, https://doi.org/10.1016/j.ygcen.2010.04.009.
Graham D E, Rechler M M, Brown A L et al. 1986. Coordinate developmental regulation of high and low molecular weight mRNAs for rat insulin-like growth factor II.Proceedings of the National Academy of Sciences of the United States of America, 83(12): 4519-4523, https://doi.org/10.1073/pnas.83.12.4519.
Greene M W, Chen T T 1997. Temporal expression pattern of insulin-like growth factor mRNA during embryonic development in a teleost, rainbow trout (Onchorynchus mykiss). Molecular Marine Biology and Biotechnology, 6(2): 144-151.
Hall T A 1999. BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symposium Series, 41: 95-98.
Jiao S, Tan X G, Wang Q et al. 2015. The olive flounder(Paralichthys olivaceus) Pax3 homologues are highly conserved, encode multiple isoforms and show unique expression patterns. Comparative Biochemistry and Physiology Part B: Biochemistry and Molecular Biology, 180: 7-15, https://doi.org/10.1016/j.cbpb.2014.10.002.
Jones J I, Clemmons D R. 1995. Insulin-like growth factors and their binding proteins: biological actions. Endocrine Reviews, 16(1): 3-34, https://doi.org/10.1210/edrv-16-1-3.
Kumar S, Stecher G, Li M et al. 2018. MEGA X: Molecular Evolutionary Genetics Analysis across computing platforms. Molecular Biology and Evolution, 35(6): 1547-1549, https://doi.org/10.1093/molbev/msy096.
Latimer A J, Appel B. 2006. Notch signaling regulates midline cell specification and proliferation in zebrafish.Developmental Biology, 298(2): 392-402, https://doi.org/10.1016/j.ydbio.2006.05.039.
Lee J E, Pintar J, Efstratiadis A. 1990. Pattern of the insulinlike growth factor II gene expression during early mouse embryogenesis. Development, 110(1): 151-159.
Li J Z, Wu P, Liu Y et al. 2014. Temporal and spatial expression of the four Igf ligands and two Igf type 1 receptors in zebrafish during early embryonic development.Gene Expression Patterns, 15(2): 104-111, https://doi.org/10.1016/j.gep.2014.05.006.
Moriyama S, Ayson F G, Kawauchi H. 2000. Growth regulation by insulin-like growth factor-I in fish. Bioscience, Biotechnology, and Biochemistry, 64(8): 1553-1562, https://doi.org/10.1271/bbb.64.1553.
Moriyama S, Yamaguchi K, Takasawa T et al. 2008.Identification of two insulin-like growth factor IIs in the Japanese eel, Anguilla japonica: cloning, tissue distribution, and expression after growth hormone treatment and seawater acclimation. Comparative Biochemistry and Physiology Part B: Biochemistry and Molecular Biology, 149(1): 47-57, https://doi.org/10.1016/j.cbpb.2007.08.005.
Nipkow M, Wirthgen E, Luft P et al. 2018. Characterization of igf1 and igf2 genes during maraena whitefish (Coregonus maraena) ontogeny and the effect of temperature on embryogenesis and igf expression. Growth Hormone& IGF Research, 40: 32-43, https://doi.org/10.1016/j.ghir.2018.04.003.
O'Dell S D, Day I N. 1998. Molecules in focus Insulin-like growth factor II (IGF-II). The International Journal of Biochemistry & Cell Biology, 30(7): 767-771, https://doi.org/10.1016/s1357-2725(98)00048-x.
Perez L, Ortiz-Delgado J B, Manchado M. 2016. Molecular characterization and transcriptional regulation by GH and GnRH of insulin-like growth factors I and II in white seabream (Diplodus sargus). Gene, 578(2): 251-262, https://doi.org/10.1016/j.gene.2015.12.030.
Perrot V, Moiseeva E B, Gozes Y et al. 1999. Ontogeny of the insulin-like growth factor system (IGF-I, IGF-II, and IGF-1R) in gilthead seabream (Sparus aurata): expression and cellular localization. General and Comparative Endocrinology, 116(3): 445-460, https://doi.org/10.1006/gcen.1999.7337.
Peterson B C, Bosworth B G, Bilodeau A L. 2005. Differential gene expression of IGF-I, IGF-II, and toll-like receptors 3 and 5 during embryogenesis in hybrid (channel×blue) and channel catfish.ComparativeBiochemistryandPhysiology Part A: Molecular & Integrative Physiology, 141(1): 42-47, https://doi.org/10.1016/j.cbpb.2005.03.009.
Pierce A L, Breves J P, Moriyama S et al. 2011. Differential regulation of Igf1 and Igf2 mRNA levels in tilapia hepatocytes: effects of insulin and cortisol on GH sensitivity. Journal of Endocrinology, 211(2): 201-210, https://doi.org/10.1530/JOE-10-0456.
Placzek M, Briscoe J. 2005. The floor plate: multiple cells, multiple signals. Nature Reviews Neuroscience, 6(3):230-240, https://doi.org/10.1038/nrn1628.
Radaelli G, Patruno M, Maccatrozzo L et al. 2003. Expression and cellular localization of insulin-like growth factor-II protein and mRNA in Sparus aurata during development.Journal of Endocrinology, 178(2): 285-299, https://doi.org/10.1677/joe.0.1780285.
Reinecke M, Björnsson B T, Dickhoff W W et al. 2005.Growth hormone and insulin-like growth factors in fish:where we are and where to go. General and Comparative Endocrinology, 142(1-2): 20-24, https://doi.org/10.1016/j.ygcen.2005.01.016.
Reinecke M. 2010. Insulin-like growth factors and fish reproduction. Biology of Reproduction, 82(4): 656-661, https://doi.org/10.1095/biolreprod.109.080093.
Rotwein P. 1991. Structure, evolution, expression and regulation of insulin-like growth factors I and II. Growth Factors, 5(1): 3-18, https://doi.org/10.3109/08977199109000267.
Soares M B, Ishii D N, Efstratiadis A. 1985. Developmental and tissue-specific expression of a family of transcripts related to rat insulin-like growth factor II mRNA.Nucleic Acids Research, 13(4): 1119-1134, https://doi.org/10.1093/nar/13.4.1119.
Sussenbach J S, Rodenburg R J T, Scheper W et al. 1993.Transcriptional and post-transcriptional regulation of the human IGF-II gene expression. In: Le Roith D, Raizada M K eds. Current Directions in Insulin-Like Growth Factor Research. Springer, Boston, p.63-71, https://doi.org/10.1007/978-1-4615-2988-0_7.
Tse M C L, Vong Q P, Cheng C H K et al. 2002. PCR-cloning and gene expression studies in common carp (Cyprinus carpio) insulin-like growth factor-II. Biochimica et Biophysica Acta, 1575(1-3): 63-74, https://doi.org/10.1016/s0167-4781(02)00244-0.
Wang B, Xu Y J, Liu X Z et al. 2019a. Molecular characterization and expression profiles of insulin-like growth factors in yellowtail kingfish (Seriola lalandi) during embryonic development. Fish Physiology and Biochemistry, 45(1):375-390, https://doi.org/10.1007/s10695-018-0570-5.
Wang C, Li X M, Dang H X et al. 2019b. Insulin-like growth factor 2 regulates the proliferation and differentiation of rat adipose-derived stromal cells via IGF-1R and IR.Cytotherapy, 21(6): 619-630, https://doi.org/10.1016/j.jcyt.2018.11.010.
White Y A, Kyle J T, Wood A W. 2009. Targeted gene knockdown in zebrafish reveals distinct intraembryonic functions for insulin-like growth factor II signaling.Endocrinology, 150(9): 4366-4375, https://doi.org/10.1210/en.2009-0356.
Wood A W, Duan C M, Bern H A. 2005. Insulin-like growth factor signaling in fish. International Review of Cytology, 243: 215-285, https://doi.org/10.1016/S0074-7696(05)43004-1.
Xu Y J, Zang K, Liu X Z et al. 2015. Insulin-like growth factors I and II in starry flounder (Platichthys stellatus): molecular cloning and differential expression during embryonic development. Fish Physiology and Biochemistry, 41(1):139-152, https://doi.org/10.1007/s10695-014-0012-y.
Zou S M, Kamei H, Modi Z et al. 2009. Zebrafish IGF genes: gene duplication, conservation and divergence, and novel roles in midline and notochord development. PLoS One, 4(9): e7026, https://doi.org/10.1371/journal.pone.0007026.