Publication: Single-cell DNA methylation sequencing reveals epigenetic alterations induced by bovine oocyte in vitro maturation
Authors
Abril-Parreño, Laura ; Lopes, Jordana S. ; Galvao, Antonio ; Kelsey, Gavin ; Coy, Pilar ; Romero Aguirregomezcorta, Jon
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Facultad de Veterinaria
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Publisher
Brazilian College of Animal Reproduction
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DOI
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info:eu-repo/semantics/lecture
Description
Abstract
Oocyte in vitro maturation (IVM) is a key step for the in vitro production of bovine embryos. However, there is a concern that this process may induce suboptimal developmental competence of bovine oocytes including epigenetic alterations. Currently, epigenetic profile of bovine oocyte IVM is relatively limited and inconsistent, probably arising from differences in the detection approach and experimental design used between studies. The aim of this work was to determine whether DNA methylome alterations are present in bovine oocytes that were in vitro matured and to identify conserved biomarkers across species. To achieve this, the results of this study were compared to those obtained in a similar study with in vitro matured porcine oocytes (unpublished data). The study was performed on 18 in vitro matured oocytes recovered from 2-8 mm follicles of abattoir-derived bovine ovaries and 28 in vivo matured oocytes collected by ovum-pick-up. Transvaginal aspiration was performed at 96–98 h after GnRH administration. The analysis of DNA methylation was performed by single-cell whole-genome bisulphite sequencing. Then, differentially methylated regions (DMRs, FDR < 0.05, FC > 0.1) were determined using the R package limma. Results showed that global DNA methylation profiles differed between in vitro and in vivo groups. Individual oocytes were clustered using Uniform Manifold Approximation and Projection analysis, which showed a clear separation within the in vivo group according to breed and age. The analysis of DMR identified a lower number of hypermethylated and hypomethylated regions in the IVM group, which were more frequent in variably methylated regions (VMRs), promoters, transcripts and imprinted genes. No differences were found in methylation of CpG islands of genes previously related to large offspring syndrome between groups. Regarding the effect on genomic imprinting, methylation was lower for IVM oocytes in the imprinted gene CDKN1C and higher in the BEGAIN gene when compared to the in vivo group. In addition, the analysis of genes that have been previously predicted for their possible function in the imprinting process showed a number of differences between the in vivo and in vitro group. For example, we identified lower methylation in in vitro matured oocytes in CpG islands of 5 “candidate” genes (SEMA7A, ZNF575, ATP4B, PDGFA, COMP) while only one was hypermethylated in the coding region of the PLCL2 gene. Finally, we identified conserved differences in methylation related to IVM between bovine and porcine oocytes for 14, 8 and 3 genomic features in the transcripts, VMRs and promoters, respectively. The findings indicate that some of the epigenetic alterations are associated with suboptimal developmental competence of IVM oocytes. In conclusion, these results could help to improve this technique when employing in vitro production procedures in cattle.
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Citation
Anim Reprod. 2024; 21(3)
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