Expression Pattern of Transcription Factor OCT4 in Early Porcine (Sus scrofa) Embryos and Its Biological Function in Trophectoderm
QI Zi-Cheng, CHAI Zhuang, CUI Hong-Di, ZHANG Yu-Ting, LIU Yan, WEI Ren-Yue, XU Qian-Qian, JIN Jun-Xue, LIU Zhong-Hua*
Key Laboratory of Animal Cellular and Genetic Engineering of Heilongjiang Province/College of Life Science, Northeast Agricultural University, Harbin 150030, China
Abstract Octamer-binding transcription factor 4 (OCT4) is a POU (Pit-Oct-Unc)-family transcription factor, which is specifically expressed in early embryos, embryonic stem cells and germ cells. Studying the expression and function of OCT4 in early porcine (Sus scrofa) embryonic development is significant for the analysis of early developmental mechanisms and pluripotency regulation. In this study, the expression pattern of OCT4 in porcine early stage embryos was studied by immunofluorescence and qPCR technology. The results showed that in parthenogenetic, in vitro fertilization and somatic cell nuclear transfer embryos, OCT4 protein widely expressed in the blastomeres from 2-cell to murola, and expressed simultaneously in the inner cell mass and trophectoderm of the blastocyst; OCT4 expression patterns in parthenogenetic and in vitro fertilization embryos were similar, the expression level declined at the 2-cell stage and increased at the 8-cell stage, and Oct4 mRNA is widely expressed in the trophoectoderm of the blastocysts. When OCT4 was specifically overexpressed in the trophoectoderm, the diameter and cell number increased significantly, trophoblast development-related genes E74-like factor 5 (ELF5) was down-regulated, EOMES was significantly up-regulated, fibroblast growth factor (FGF) signaling pathway related genes were significantly down-regulated and polarization-related genes PKCα (protein kinase Cα) and ERZIN were significantly down-regulated, methylation gene TET1 was significantly up-regulated, and DNA methyltransferases 3B (DNMT3b) was significantly down-regulated. The results indicate that OCT4 keeps expressing in the trophectoderm and might regulate proliferation after the first lineage differentiation. This study provides basic material for further study of porcine early embryonic development mechanisms and the establishment of porcine embryonic stem cells.
QI Zi-Cheng,CHAI Zhuang,CUI Hong-Di, et al. Expression Pattern of Transcription Factor OCT4 in Early Porcine (Sus scrofa) Embryos and Its Biological Function in Trophectoderm[J]. 农业生物技术学报, 2021, 29(10): 1855-1868.
[1] 伟人悦. 2020. 猪诱导多能性干细胞定向分化为血管内皮细胞的研究[D]. 博士学位论文, 东北农业大学, 导师: 刘忠华, pp. 32-33. (Wei R Y.2020. Differentiation of porcine induced pluripotent stem cells into endothelial cells[D]. Thesis for Ph.D., Northeast Agricultural University, Supevisor: Liu Z H, pp. 32-33.) [2] Bou G, Liu S, Guo J, et al.2016. Cdx2 represses Oct4 function via inducing its proteasome-dependent degradation in early porcine embryos[J]. Developmental Biology, 410(1): 36-44. [3] Bou G, Liu S, Sun M, et al.2017. CDX2 is essential for cell proliferation and polarity in porcine blastocysts[J]. Development, 144(7): 1296-1306. [4] Cauffman G, van de Velde H, Liebaers I, et al.2005. Oct-4 mRNA and protein expression during human preimplantation development[J]. Molecular Human Reproduction, 11(3): 173-181. [5] Chrysanthou S, Senner C E, Woods L, et al.2018. A critical role of TET1/2 proteins in cell-cycle progression of trophoblast stem cells[J]. Stem Cell Reports, 10(4): 1355-1368. [6] Daigneault B W, Rajput S, Smith G W, et al.2018. Embryonic POU5F1 is required for expanded bovine blastocyst formation[J]. Scientific Reports, 8(1): 7753. [7] Degrelle S A, Campion E, Cabau C, et al.2005. Molecular evidence for a critical period in mural trophoblast development in bovine blastocysts[J]. Developmental Biology, 288(2): 448-460. [8] Fogarty N M E, McCarthy A, Snijders K E, et al.2017. Genome editing reveals a role for OCT4 in human embryogenesis[J]. Nature, 550(7674): 67-73. [9] Frum T, Halbisen M A, Wang C, et al.2013. Oct4 cell-autonomously promotes primitive endoderm development in the mouse blastocyst[J]. Developmental Cell, 25(6): 610-622. [10] Gao Y, Jammes H, Rasmussen M A, et al.2011. Epigenetic regulation of gene expression in porcine epiblast, hypoblast, trophectoderm and epiblast-derived neural progenitor cells[J]. Epigenetics, 6(9): 1149-1161. [11] Hambiliki F, Strom S, Zhang P, et al.2012. Co-localization of NANOG and OCT4 in human pre-implantation embryos and in human embryonic stem cells[J]. Journal of Assisted Reproduction and Genetics, 29(10): 1021-1028. [12] Hancock S N, Agulnik S I, Silver L M, et al.1999. Mapping and expression analysis of the mouse ortholog of xenopus eomesodermin[J]. Mechanisms of Development, 81(1-2): 205-208. [13] Harris D, Huang B, Oback B.2013. Inhibition of MAP2K and GSK3 signaling promotes bovine blastocyst development and epiblast-associated expression of pluripotency factors[J]. Biology of Reproduction, 88(3): 74. [14] He S, Pant D, Schiffmacher A, et al.2006. Developmental expression of pluripotency determining factors in caprine embryos: Novel pattern of NANOG protein localization in the nucleolus[J]. Molecular Reproduction and Development, 73(12): 1512-1522. [15] He Y F, Li B Z, Li Z, et al.2011. Tet-mediated formation of 5-carboxylcytosine and its excision by TDG in mammalian DNA[J]. Science, 333(6047): 1303-1307. [16] Jerabek S, Merino F, Scholer H R, et al.2014. OCT4: Dynamic DNA binding pioneers stem cell pluripotency[J]. Biochimica et Biophysica Acta, 1839(3): 138-154. [17] Johnson M H, Ziomek C A.1981. Induction of polarity in mouse 8-cell blastomeres: Specificity, geometry, and stability[J]. The Journal of Cell Biology, 91(1): 303-308. [18] Kim S J, Koo O J, Park H J, et al.2015. Oct4 overexpression facilitates proliferation of porcine fibroblasts and development of cloned embryos[J]. Zygote, 23(5): 704-711. [19] Kirchhof N, Carnwath J W, Lemme E, et al.2000. Expression pattern of Oct-4 in preimplantation embryos of different species[J]. Biology of Reproduction, 63(6): 1698-1705. [20] Kuijk E W, Du Puy L, van Tol H T, et al.2008. Differences in early lineage segregation between mammals[J]. Developmental Dynamics, 237(4): 918-927. [21] Kurowski A, Molotkov A, Soriano P.2019. FGFR1 regulates trophectoderm development and facilitates blastocyst implantation[J]. Developmental Biology, 446(1): 94-101. [22] Le Bin G C, Munoz-Descalzo S, Kurowski A, et al.2014. Oct4 is required for lineage priming in the developing inner cell mass of the mouse blastocyst[J]. Development, 141(5): 1001-1010. [23] Li L Y, Li M M, Yang S F, et al.2016. Inhibition of FGF signalling pathway augments the expression of pluripotency and trophoblast lineage marker genes in porcine parthenogenetic blastocyst[J]. Reproduction in Domestic Animals, 51(5): 649-656. [24] Liu S, Bou G, Sun R, et al.2015. Sox2 is the faithful marker for pluripotency in pig: Evidence from embryonic studies[J]. Developmental Dynamics, 244(4): 619-627. [25] Maitre J L, Turlier H, Illukkumbura R, et al.2016. Asymmetric division of contractile domains couples cell positioning and fate specification[J]. Nature, 536(7616): 344-348. [26] Malashicheva A, Kanzler B, Tolkunova E, et al.2007. Lentivirus as a tool for lineage-specific gene manipulations[J]. Genesis, 45(7): 456-459. [27] McLean Z, Meng F, Henderson H, et al.2014. Increased MAP kinase inhibition enhances epiblast-specific gene expression in bovine blastocysts[J]. Biology of Reproduction, 91(2): 49. [28] Nichols J, Silva J, Roode M, et al.2009. Suppression of Erk signalling promotes ground state pluripotency in the mouse embryo[J]. Development, 136(19): 3215-3222. [29] Nichols J, Zevnik B, Anastassiadis K, et al.1998. Formation of pluripotent stem cells in the mammalian embryo depends on the POU transcription factor Oct4[J]. Cell, 95(3): 379-391. [30] Nishioka N, Inoue K, Adachi K, et al.2009. The Hippo signaling pathway components Lats and Yap pattern Tead4 activity to distinguish mouse trophectoderm from inner cell mass[J]. Developmental Cell, 16(3): 398-410. [31] Niwa H, Toyooka Y, Shimosato D, et al.2005. Interaction between Oct3/4 and Cdx2 determines trophectoderm differentiation[J]. Cell, 123(5): 917-929. [32] Olariu V, Lövkvist C, Sneppen K.2016. Nanog, Oct4 and Tet1 interplay in establishing pluripotency[J]. Scientific Reports, 6: 25438. [33] Pearton D J, Broadhurst R, Donnison M, et al.2011. Elf5 regulation in the trophectoderm[J]. Developmental Biology, 360(2): 343-350. [34] Pesce M, Scholer H R.2001. Oct-4: Gatekeeper in the beginnings of mammalian development[J]. Stem Cells, 19(4): 271-278. [35] Pratt H P, Ziomek C A, Reeve W J, et al.1982. Compaction of the mouse embryo: An analysis of its components[J]. Journal of Embryology and Experimental Morphology, 70: 113-132. [36] Rosner M H, Vigano M A, Ozato K, et al.1990. A POU-domain transcription factor in early stem cells and germ cells of the mammalian embryo[J]. Nature, 345(6277): 686-692. [37] Russ A P, Wattler S, Colledge W H, et al.2000. Eomesodermin is required for mouse trophoblast development and mesoderm formation[J]. Nature, 404(6773): 95-99. [38] Scholer H R, Dressler G R, Balling R, et al.1990a. Oct-4: A germline-specific transcription factor mapping to the mouse t-complex[J]. The EMBO Journal, 9(7): 2185-2195. [39] Scholer H R, Ruppert S, Suzuki N, et al.1990b. New type of POU domain in germ line-specific protein Oct-4[J]. Nature, 344(6265): 435-439. [40] Simmet K, Zakhartchenko V, Philippou-Massier J, et al.2018. OCT4/POU5F1 is required for NANOG expression in bovine blastocysts[J]. Proceedings of the National Academy of Sciences of the USA, 115(11): 2770-2775. [41] Strumpf D, Mao C A, Yamanaka Y, et al.2005. Cdx2 is required for correct cell fate specification and differentiation of trophectoderm in the mouse blastocyst[J]. Development, 132(9): 2093-2102. [42] Yeom Y I, Fuhrmann G, Ovitt C E, et al.1996. Germline regulatory element of Oct-4 specific for the totipotent cycle of embryonal cells[J]. Development, 122(3): 881-894. [43] Zhu J, Wang K, Li T, et al.2017. Hypoxia-induced TET1 facilitates trophoblast cell migration and invasion through HIF1α signaling pathway[J]. Scientific Reports, 7(1): 8077.
