YTHDF2 Regulates Maternal Transcriptome Degradation and Embryo Development in Goat

Molecular Therapy - Nucleic Acids

Wan

Chen

et al. 2021

Self Cite

Long non-coding RNAs (lncRNAs) are involved in shaping chromosome conformation and regulation of preimplantation development. However, the role of lncRNA during somatic cell nuclear transfer (SCNT) reprogramming remains largely unknown. In the present study, we identified 114 upregulated lncRNAs in the 8-cell SCNT embryos as candidate key molecules involved in nuclear reprogramming in goat. We found that H3K4me3 was an epigenetic barrier in goat nuclear reprogramming that and injection of Kdm5b mRNA greatly improved SCNT embryos development through removal of H3K4me3. We further reported that knockdown of lnc_3712 increased the expression of Kdm5b, which led to H3K4me3 demethylation. Of note, the development of goat SCNT embryos was improved when lnc_3712 was knocked down, whereas the blastocyst rate showed no difference in lnc_3712 and Kdm5b double knockdown SCNT embryos compared with the negative control SCNT embryos. Specifically, in lnc_3712 knockdown SCNT embryos, partial of the transcriptional activity and the expression of critical embryonic genes ( Wee1 , Ctsb , and Ybx1 ) were similar with that of in vitro fertilization embryos. Therefore, our results elucidate the critical role of lnc_3712 in regulating the development of goat SCNT embryos via repressing Kdm5b, which advances our current understanding of the role of lncRNAs during nuclear reprogramming.

Section: Discussionmentioning

confidence: 83%

Long non-coding RNA lnc_3712 impedes nuclear reprogramming via repressing Kdm5b

Molecular Therapy - Nucleic Acids

Wan

Chen

et al. 2021

Self Cite

“…mRNA stability was also regulated by m6A. The m6A reader IGF2BP1 and YTHDF2 have been reported to regulate the early embryo development [9,10,29], while the m6A methyltransferase METTL3 was essential for fertility [30]. The down-regulation of METTL3, IGF2BP1, and YTHDF2 might impede the early embryo development.…”

Section: Discussionmentioning

confidence: 99%

“…In vitro maturation (IVM) was performed as previously described [37]. Brie y, cumulus-oocyte complexes (COCs) with more than two layers of compact cumulus cells and a dense, homogeneous cytoplasm were obtained and cultured in groups of 20 in 60-μL droplets of IVM medium at 38.5°C, with 5% CO 2 , 95% air, and saturated humidity, for 20-22 h. Subsequently, in vitro fertilization (IVF) was performed as previously described [10]. After culturing for 16 h at 38.5°C, with 5% CO 2 , 5% O 2 , 90% N 2 , and saturated humidity, the zygotes were collected for knockdown experiment.…”

Section: In Vitro Fertilizationmentioning

confidence: 99%

“…For example, B-cell Translocation Gene-4 (BTG4) bridged CNOT to EIF4E, and facilitated decay of maternal mRNA [8]. De ciency of m6A reader protein YTH N6-methyladenosine RNA binding protein 2 (YTHDF2) decelerated the decay of m6A-modi ed maternal mRNAs and impeded ZGA initiation [9,10]. These studies suggesting that maternal factors play pivotal roles during MZT.…”

Section: Introductionmentioning

confidence: 99%

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YBX1 Mediates Alternative Splicing and Maternal mRNA Decay During Pre-Implantation Development

Chen

Liu

et al. 2021

Preprint

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Background: In mammals, maternal gene products decay and zygotic genome activation (ZGA) during maternal to zygotic transition (MZT) is critical for pre-implantation. Y-box binding protein YBX1 plays vital roles in RNA stabilization and transcriptional regulation, but its roles in pre-implantation development remain to be elucidated. The objective of this study is to investigate the role and the molecular mechanisms of YBX1 during MZT.Methods: RNA-seq datasets in mice, human, bovine, and goat embryos were re-analyzed. YBX1 was knocked down by siRNA microinjection. The 8-cell stage embryos were collected for RNA-seq. The differentially expressed genes and alternative splicing (AS) events were identified using DESeq2 and rMATs, respectively. GO/KEGG/GSEA enrichment analysis was performed using clusterProfiler and enrichplot. Furthermore, 5-EU staining was performed to confirm the effect of YBX1 knockdown on transcriptional activity.Results: The expression of YBX1 was increased during MZT in goat, bovine, human, and mice. By microinjection of siRNA against YBX1, we successfully knocked down YBX1, and the embryo development was impaired in YBX1 knockdown embryos. Using RNA-seq, we identified 1623 up-regulated and 3531 down-regulated genes in the 8-cell stage YBX1 knockdown embryos. Of note, the down-regulated genes were enriched in regulation of RNA/mRNA stability and spliceosome, suggesting that YBX1 might medicate RNA stability and AS. To this end, we identified 3284 differential AS events and 1322 differentially expressed maternal mRNAs at the 8-cell stage YBX1 knockdown embryos. Meanwhile, the splicing factors and mRNA decay related showed aberrant expression. Moreover, the transcriptional activity during ZGA in goat and mice was compromised when YBX1 was knocked down.Conclusion: Our results identify that YBX1 serves an important role in maternal mRNA decay, alternative splicing, and the transcriptional activity required for early embryogenesis, which will broaden the current understanding of YBX1 functions during the stochastic reprogramming events.

“…GO function and KEGG pathway enrichment analyses were performed with reference to previous studies [ 23 ]. GO and KEGG terms with FDR adjusted p < 0.05 were considered statistically significant.…”

Section: Methodsmentioning

confidence: 99%

Comprehensive Transcriptome Analysis of mRNA Expression Patterns of Early Embryo Development in Goat under Hypoxic and Normoxic Conditions

Wan

et al. 2021

Biology

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It has been reported that hypoxic environments were more suitable for the in vitro development of mammalian embryos, but the underlying mechanisms were still unclear. In the present study, RNA-seq was performed to compare 8-cell-stage and blastocyst-stage goat embryos under hypoxic and normoxic conditions; zygotes were checked at 72 and 168 h to 8-cell stage (L8C) and blastocyst stage (LM) in hypoxic conditions and 8-cell stage (H8C) and blastocyst stage (HM) in normoxic conditions. In the H8C and L8C groups, 399 DEGs were identified, including 348 up- and 51 down-regulated DEGs. In the HM and LM groups, 1710 DEGs were identified, including 1516 up- and 194 down-regulated DEGs. The expression levels of zygotic genes, transcription factors, and maternal genes, such as WEE2, GDF9, HSP70.1, BTG4, and UBE2S showed significant changes. Functional enrichment analysis indicated that these DEGs were mainly related to biological processes and function regulation. In addition, combined with the pathway–gene interaction network and protein–protein interaction network, twenty-two of the hub genes were identified and they are mainly involved in energy metabolism, immune stress response, cell cycle, receptor binding, and signal transduction pathways. The present study provides comprehensive insights into the effects of oxidative stress on early embryo development in goats.