Differential expression profiles of long non‑coding RNAs during the mouse pronuclear stage under normal gravity and simulated microgravity

Feng,Meiying; Dang,Nannan; Bai,Yinshan; Wei,Hengxi; Meng,Li; Wang,Kai; Zhao,Zhihong; Chen,Yun; Gao,Fenglei; Chen,Zhilin; Li,Li; Zhang,Shouquan

doi:10.3892/mmr.2018.9675

Differential expression profiles of long non‑coding RNAs during the mouse pronuclear stage under normal gravity and simulated microgravity

Authors:
- Meiying Feng
- Nannan Dang
- Yinshan Bai
- Hengxi Wei
- Li Meng
- Kai Wang
- Zhihong Zhao
- Yun Chen
- Fenglei Gao
- Zhilin Chen
- Li Li
- Shouquan Zhang
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Affiliations: College of Animal Science, National Engineering Research Center for Breeding Swine Industry, Guangdong Provincial Key Lab of Agro‑Animal Genomics and Molecular Breeding, College of Animal Science, South China Agricultural University, Guangzhou, Guangdong 510642, P.R. China
Published online on: November 20, 2018 https://doi.org/10.3892/mmr.2018.9675
Pages: 155-164
Copyright: © Feng et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Pronuclear migration, which is the initial stage of embryonic development and the marker of zygote formation, is a crucial process during mammalian preimplantation embryonic development. Recent studies have revealed that long non‑coding RNAs (lncRNAs) serve an important role in early embryonic development. However, the functional regulation of lncRNAs in this process has yet to be elucidated, largely due to the difficulty of assessing gene expression alterations during the very short time in which pronuclear migration occurs. It has previously been reported that migration of the pronucleus of a zygote can be obstructed by simulated microgravity. To investigate pronuclear migration in mice, a rotary cell culture system was employed, which generates simulated microgravity, in order to interfere with murine pronuclear migration. Subsequently, lncRNA sequencing was performed to investigate the mechanism underlying this process. In the present study, a comprehensive analysis of lncRNA profile during the mouse pronuclear stage was conducted, in which 3,307 lncRNAs were identified based on single‑cell RNA sequencing data. Furthermore, 52 lncRNAs were identified that were significantly differentially expressed. Subsequently, 10 lncRNAs were selected for validation by reverse transcription‑quantitative polymerase chain reaction, in which the same relative expression pattern was observed. The results revealed that 12 lncRNAs (lnc006745, lnc007956, lnc013100, lnc013782, lnc017097, lnc019869, lnc025838, lnc027046, lnc005454, lnc007956, lnc019410 and lnc019607), with tubulin β 4B class IVb or actinin α 4 as target genes, may be associated with the expression of microtubule and microfilament proteins. Binding association was confirmed using a dual‑luciferase reporter assay. Finally, Gene Ontology analysis revealed that the target genes of the differentially expressed lncRNAs participated in cellular processes associated with protein transport, binding, catalytic activity, membrane‑bounded organelle, protein complex and the cortical cytoskeleton. These findings suggested that these lncRNAs may be associated with migration of the mouse pronucleus.

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1	Aoki F, Worrad DM and Schultz RM: Regulation of transcriptional activity during the first and second cell cycles in the preimplantation mouse embryo. Dev Biol. 181:296–307. 1997. View Article : Google Scholar : PubMed/NCBI
2	Latham KE, Garrels JI, Chang C and Solter D: Quantitative analysis of protein synthesis in mouse embryos. I. Extensive reprogramming at the one- and two-cell stages. Development. 112:921–932. 1991.PubMed/NCBI
3	Gundersen GG and Worman HJ: Nuclear positioning. Cell. 152:1376–1389. 2013. View Article : Google Scholar : PubMed/NCBI
4	Clift D and Schuh M: Restarting life: Fertilization and the transition from meiosis to mitosis. Nat Rev Mol Cell Biol. 14:549–562. 2013. View Article : Google Scholar : PubMed/NCBI
5	Schatten G, Simerly C and Schatten H: Microtubule configurations during fertilization, mitosis, and early development in the mouse and the requirement for egg microtubule-mediated motility during mammalian fertilization. Proc Natl Acad Sci USA. 82:4152–4156. 1985. View Article : Google Scholar : PubMed/NCBI
6	Wu C, Guo X, Wang F, Li X, Tian XC, Li L, Wu Z and Zhang S: Simulated microgravity compromises mouse oocyte maturation by disrupting meiotic spindle organization and inducing cytoplasmic blebbing. PLoS One. 6:e222142011. View Article : Google Scholar : PubMed/NCBI
7	Kutter C, Watt S, Stefflova K, Wilson MD, Goncalves A, Ponting CP, Odom DT and Marques AC: Rapid turnover of long noncoding RNAs and the evolution of gene expression. PLoS Genet. 8:e10028412012. View Article : Google Scholar : PubMed/NCBI
8	Hamazaki N, Uesaka M, Nakashima K, Agata K and Imamura T: Gene activation-associated long noncoding RNAs function in mouse preimplantation development. Development. 142:910–920. 2015. View Article : Google Scholar : PubMed/NCBI
9	Zhang K, Huang K, Luo Y and Li S: Identification and functional analysis of long non-coding RNAs in mouse cleavage stage embryonic development based on single cell transcriptome data. BMC Genomics. 15:8452014. View Article : Google Scholar : PubMed/NCBI
10	Yan L, Yang M, Guo H, Yang L, Wu J, Li R, Liu P, Lian Y, Zheng X, Yan J, et al: Single-cell RNA-Seq profiling of human preimplantation embryos and embryonic stem cells. Nat Struct Mol Biol. 20:1131–1139. 2013. View Article : Google Scholar : PubMed/NCBI
11	Wang KC and Chang HY: Molecular mechanisms of long noncoding RNAs. Mol Cell. 43:904–914. 2011. View Article : Google Scholar : PubMed/NCBI
12	Mercer TR, Dinger ME and Mattick JS: Long non-coding RNAs: Insights into functions. Nat Rev Genet. 10:155–159. 2009. View Article : Google Scholar : PubMed/NCBI
13	Mercer TR and Mattick JS: Structure and function of long noncoding RNAs in epigenetic regulation. Nat Struct Mol Biol. 20:300–307. 2013. View Article : Google Scholar : PubMed/NCBI
14	Ulitsky I and Bartel DP: lincRNAs: Genomics, evolution, and mechanisms. Cell. 154:26–46. 2013. View Article : Google Scholar : PubMed/NCBI
15	Pauli A, Rinn JL and Schier AF: Non-coding RNAs as regulators of embryogenesis. Nat Rev Genet. 12:136–149. 2011. View Article : Google Scholar : PubMed/NCBI
16	Hamatani T, Carter MG, Sharov AA and Ko MS: Dynamics of global gene expression changes during mouse preimplantation development. Dev Cell. 6:117–131. 2004. View Article : Google Scholar : PubMed/NCBI
17	Wang QT, Piotrowska K, Ciemerych MA, Milenkovic L, Scott MP, Davis RW and Zernicka-Goetz M: A genome-wide study of gene activity reveals developmental signaling pathways in the preimplantation mouse embryo. Dev Cell. 6:133–144. 2004. View Article : Google Scholar : PubMed/NCBI
18	Xie D, Chen CC, Ptaszek LM, Xiao S, Cao X, Fang F, Ng HH, Lewin HA, Cowan C and Zhong S: Rewirable gene regulatory networks in the preimplantation embryonic development of three mammalian species. Genome Res. 20:804–815. 2010. View Article : Google Scholar : PubMed/NCBI
19	Tang F, Barbacioru C, Nordman E, Bao S, Lee C, Wang X, Tuch BB, Heard E, Lao K and Surani MA: Deterministic and stochastic allele specific gene expression in single mouse blastomeres. PLoS One. 6:e212082011. View Article : Google Scholar : PubMed/NCBI
20	Li XY, Cui XS and Kim NH: Transcription profile during maternal to zygotic transition in the mouse embryo. Reprod Fertil Dev. 18:635–645. 2006. View Article : Google Scholar : PubMed/NCBI
21	Tang F, Barbacioru C, Wang Y, Nordman E, Lee C, Xu N, Wang X, Bodeau J, Tuch BB, Siddiqui A, et al: mRNA-Seq whole-transcriptome analysis of a single cell. Nat Methods. 6:377–382. 2009. View Article : Google Scholar : PubMed/NCBI
22	Fan X, Zhang X, Wu X, Guo H, Hu Y, Tang F and Huang Y: Single-cell RNA-seq transcriptome analysis of linear and circular RNAs in mouse preimplantation embryos. Genome Biol. 16:1482015. View Article : Google Scholar : PubMed/NCBI
23	Serra L, Chang DZ, Macchietto M, Williams K, Murad R, Lu D, Dillman AR and Mortazavi A: Adapting the smart-seq2 protocol for robust single worm RNA-seq. Bio Protoc. 8:e27292018. View Article : Google Scholar : PubMed/NCBI
24	Trapnell C, Roberts A, Goff L, Pertea G, Kim D, Kelley DR, Pimentel H, Salzberg SL, Rinn JL and Pachter L: Differential gene and transcript expression analysis of RNA-seq experiments with TopHat and Cufflinks. Nat Protoc. 7:562–578. 2012. View Article : Google Scholar : PubMed/NCBI
25	Kong L, Zhang Y, Ye ZQ, Liu XQ, Zhao SQ, Wei L and Gao G: CPC: Assess the protein-coding potential of transcripts using sequence features and support vector machine. Nucleic Acids Res. 35:W345–W349. 2007. View Article : Google Scholar : PubMed/NCBI
26	Love MI, Huber W and Anders S: Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol. 15:5502014. View Article : Google Scholar : PubMed/NCBI
27	Livak KJ and Scmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods. 25:402–408. 2001. View Article : Google Scholar : PubMed/NCBI
28	Shannon P, Markiel A, Ozier O, Baliga NS, Wang JT, Ramage D, Amin N, Schwikowski B and Ideker T: Cytoscape: A software environment for integrated models of biomolecular interaction networks. Genome Res. 13:2498–2504. 2003. View Article : Google Scholar : PubMed/NCBI
29	Wu GQ, Wang X, Zhou HY, Chai KQ, Xue Q, Zheng AH, Zhu XM, Xiao JY, Ying XH, Wang FW, et al: Evidence for transcriptional interference in a dual-luciferase reporter system. Sci Rep. 5:176752015. View Article : Google Scholar : PubMed/NCBI
30	Tafer H and Hofacker IL: RNAplex: A fast tool for RNA-RNA interaction search. Bioinformatics. 24:2657–2663. 2008. View Article : Google Scholar : PubMed/NCBI
31	Cote I, Vigneault C, Laflamme I, Laquerre J, Fournier E, Gilbert I, Scantland S, Gagne D, Blondin P and Robert C: Comprehensive cross production system assessment of the impact of in vitro microenvironment on the expression of messengers and long non-coding RNAs in the bovine blastocyst. Reproduction. 142:99–112. 2011. View Article : Google Scholar : PubMed/NCBI
32	Plourde D, Vigneault C, Lemay A, Breton L, Gagne D, Laflamme I, Blondin P and Robert C: Contribution of oocyte source and culture conditions to phenotypic and transcriptomic variation in commercially produced bovine blastocysts. Theriogenology. 78:116–131. 2012. View Article : Google Scholar : PubMed/NCBI
33	Shalek AK, Satija R, Adiconis X, Gertner RS, Gaublomme JT, Raychowdhury R, Schwartz S, Yosef N, Malboeuf C, Lu D, et al: Single-cell transcriptomics reveals bimodality in expression and splicing in immune cells. Nature. 498:236–240. 2013. View Article : Google Scholar : PubMed/NCBI
34	Wills QF, Livak KJ, Tipping AJ, Enver T, Goldson AJ, Sexton DW and Holmes C: Single-cell gene expression analysis reveals genetic associations masked in whole-tissue experiments. Nat Biotechnol. 31:748–752. 2013. View Article : Google Scholar : PubMed/NCBI
35	Wuhr M, Dumont S, Groen AC, Needleman DJ and Mitchison TJ: How does a millimeter-sized cell find its center? Cell Cycle. 8:1115–1121. 2009. View Article : Google Scholar : PubMed/NCBI
36	Chew TG, Lorthongpanich C, Ang WX, Knowles BB and Solter D: Symmetric cell division of the mouse zygote requires an actin network. Cytoskeleton (Hoboken). 69:1040–1046. 2012. View Article : Google Scholar : PubMed/NCBI
37	Reinsch S and Gonczy P: Mechanisms of nuclear positioning. J Cell Sci. 111:2283–2295. 1998.PubMed/NCBI
38	Gadadhar S, Bodakuntla S, Natarajan K and Janke C: The tubulin code at a glance. J Cell Sci. 130:1347–1353. 2017. View Article : Google Scholar : PubMed/NCBI
39	Sarkar H, Arya S, Rai U and Majumdar SS: A study of differential expression of testicular genes in various reproductive phases of hemidactylus flaviviridis (Wall Lizard) to derive their association with onset of spermatogenesis and its relevance to mammals. PLoS One. 11:e1511502016. View Article : Google Scholar
40	Thomas DG and Robinson DN: The fifth sense: Mechanosensory regulation of alpha-actinin-4 and its relevance for cancer metastasis. Semin Cell Dev Biol. 71:68–74. 2017. View Article : Google Scholar : PubMed/NCBI
41	Zhao X, Khurana S, Charkraborty S, Tian Y, Sedor JR, Bruggman LA and Kao HY: α Actinin 4 (ACTN4) regulates glucocorticoid receptor-mediated transactivation and transrepression in podocytes. J Biol Chem. 292:1637–1647. 2017. View Article : Google Scholar : PubMed/NCBI
42	Hsu KS and Kao HY: Alpha-actinin 4 and tumorigenesis of breast cancer. Vitam Horm. 93:323–351. 2013. View Article : Google Scholar : PubMed/NCBI
43	Shao H, Wang JH, Pollak MR and Wells A: α-actinin-4 is essential for maintaining the spreading, motility and contractility of fibroblasts. PLoS One. 5:e139212010. View Article : Google Scholar : PubMed/NCBI
44	Hamill KJ, Hopkinson SB, Skalli O and Jones JC: Actinin-4 in keratinocytes regulates motility via an effect on lamellipodia stability and matrix adhesions. FASEB J. 27:546–556. 2013. View Article : Google Scholar : PubMed/NCBI