62 Transcript profiling of haploid bovine embryos during embryonic genome activation
L. Águila A B , R. Perecin A C , J. Therrien A , R. Sampaio A , F. Meirelles C , R. Felmer B and L. Smith AA Centre de Recherche en Reproduction et Fértilité (CRRF), Université de Montréal, St-Hyacinthe, Canada
B Laboratory of Reproduction, Centre of Reproductive Biotechnology (CEBIOR-BIOREN), Faculty of Agriculture and Forestry, Universidad de La Frontera, Temuco, Araucania, Chile
C Department of Veterinary Medicine, Faculty of Animal Sciences and Food Engineering, University of Sao Paulo, Pirassununga, São Paulo, Brazil
Reproduction, Fertility and Development 35(2) 157-157 https://doi.org/10.1071/RDv35n2Ab62
Published: 5 December 2022
© 2023 The Author(s) (or their employer(s)). Published by CSIRO Publishing on behalf of the IETS
Haploid embryonic stem cell lines (ESC) obtained from haploid androgenetic embryos (HAE) have been obtained in mice and humans. However, most bovine HAE undergo developmental arrest before reaching the blastocyst stage, which precludes their use for deriving ESC. To further our understanding of factors involved in the developmental features of bovine HAE, we performed a comprehensive comparison of the transcriptomic profile during the major phase of embryonic genome activation (EGA). Groups of haploid androgenetic (HA), haploid parthenogenetic, and biparental (BI) 8-cell stage embryos were produced using, respectively, intracytoplasmic sperm injection (ICSI), cycloheximide activation, and spindle enucleation after ICSI. Alfa-amanitin (AA) was used to identify oocyte-derived transcripts. The transcriptome from 5 biological replicates generated 125 Gb of data. The mapping rate was above 85% using STAR with bovine reference genome ARS-UCD1.2. After quality control, we detected a total of 19,558 expressed genes (minimum of 1 count/sample,16 counts/gene, in at least 70% of samples/group). To determine differentially expressed genes (DEGs) we used DESEqn 2 (padj < 0.05 and log2 fold change > 0.5). Average Pearson correlation showed a great correlation between samples, with R values of 0.87, 0.89, 0.91, 0.91, 0.85, and 0.93, respectively, for HP, HP+AA, BI, BI+AA, HA, and HA+AA (global R-value of 0.76). Principal component analysis (PCA) revealed closer clustering between HP and BI. In contrast, HA embryos scattered differentially from the other groups. We found 2,867 DEGs between HA and BI, and 1,823 DEGs between HA and HP. However, only 196 DEGs between HP and BI. Co-expression analysis detected a total of 675 DEGs between HA and BI, 718 genes between BI and HP, and only 207 from HP and HA. Additionally, there were no important differences between AA groups, but 7,809, 6,195, and 5,240 DEGs were found between BI vs BI+AA, HA vs AA+HA, and HP vs AA+HP, respectively. Among the top hundred DEGs, transcripts related to metabolism (TMEM250, SLC52A2, ADCY7), phosphorylation (CAMKK2), transcriptional regulation (HDAC7, ZNF821, L3MBTL2), and cell growth (NXN, CCDC85B) showed a differential pattern between haploid and biparental samples. In addition, the imprinted genes SNRPN and SNURF were highly represented in HA and BI compared to HP. Similarly, the PEG3 gene was more represented in HP than BI. Furthermore, the human imprinted genes CDH18 and SVOPL were more expressed in HA and HP, respectively. These data suggest that unbalanced transcript abundance during EGA and major transcriptomic differences in non-imprinted and imprinted genes might be behind developmental constraints of bovine haploid embryos. In addition, these data uncover potential unreported imprinted genes for the bovine species.
This research was supported by a grant from NSERC-Canada with Boviteq inc. (CRDPJ 536636-18 and CRDPJ 487107-45 to LS) and a scholarship by ANID-Chile POSTDOCTORADO BECAS CHILE/2017 – 74180059 and Programa de Formacion de Investigadores Postdoctorales, Universidad de La Frontera (PDT21-0001) (LA).
