Coordination of zygotic genome activation entry and exit by H3K4me3 and H3K27me3 in porcine early embryos

Abstract

Histone modifications are critical epigenetic indicators of chromatin state associated with gene expression. Although the reprogramming patterns of H3K4me3 and H3K27me3 have been elucidated in mouse and human preimplantation embryos, the relationship between these marks and zygotic genome activation (ZGA) remains poorly understood. By ultra-low-input native chromatin immunoprecipitation and sequencing, we profiled global H3K4me3 and H3K27me3 in porcine oocytes and in vitro fertilized (IVF) embryos. We found that promoters of ZGA genes occupied sharp H3K4me3 peaks in oocytes, and these peaks became broader after fertilization, and reshaped into sharp again during ZGA. By simultaneous depletion of H3K4me3 demethylase KDM5B and KDM5C, we determined that broad H3K4me3 domain maintenance impaired ZGA gene expression, suggesting its function to prevent premature ZGA entry. By contrast, broad H3K27me3 domains underwent global removal upon fertilization, followed by a re-establishment for H3K4me3/H3K27me3 bivalency in morulae. We also found that bivalent marks were deposited at promoters of ZGA genes, and inhibiting this deposition was correlated with the activation of ZGA genes. It suggests that promoter bivalency contributes to ZGA exit in porcine embryos. Moreover, we demonstrated that aberrant reprogramming of H3K4me3 and H3K27me3 triggered ZGA dysregulation in somatic cell nuclear transfer (SCNT) embryos, whereas H3K27me3-mediated imprinting did not exist in porcine IVF and SCNT embryos. Our findings highlight two previously unknown epigenetic reprogramming modes coordinated with ZGA in porcine preimplantation embryos. Finally, the similarities observed between porcine and human histone modification dynamics suggest that the porcine embryo may also be a useful model for human embryo research.

Epigenetic-related transcriptional reprogramming elucidated by identification and validation of a novel reference gene combination for RT-qPCR studies in porcine oocytes of contrasting quality

“…Demethylation of H3K4me3 by KDM5 proteins is crucial for proper EGA [ 9 , 12 , 16 ]. Of the KDM5 family, only lysine demethylase 5A ( KDM5A ) transcripts were detected in porcine oocytes [ 9 ]. These transcripts are stored during the oocyte growth phase [ 1 , 2 ].…”

“…The expression stability of five references genes, previously validated and/or utilised as reference genes in similar studies; Actin beta ( ACTB ) [ 18 , 31 – 33 ], glyceraldehyde-3-phosphate dehydrogenase ( GAPDH ) [ 9 , 18 , 31 , 34 ], hypoxanthine phosphoribosyltransferase 1 ( HPRT1 ) [ 35 ], tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein gamma ( YWHAG ) [ 18 ] and hydroxymethylbilane synthase ( HMBS aka PBGD ) [ 36 , 37 ], were compared employing the software program NormFinder [ 23 ]. GeNorm [ 22 ] is perhaps the most widely used algorithm for assessing reference gene stability.…”

“…In porcine oocytes, the growth phase lasts until the oocyte is enclosed in a follicle of approximately 2–3 mm in diameter, at which point transcription in the germinal vesicle nucleus is silenced prior to meiotic resumption [ 3 – 6 ]. Transcript abundance will subsequently gradually decline, as the oocyte relies on post-transcriptional regulation for selective degradation and usage until embryonic genome activation (EGA) [ 4 , 5 , 7 , 8 ], which occurs around the 4-cell stage in porcine embryos [ 9 , 10 ]. Gene expression in oocytes therefore represents a peculiar case where it does not merely reflect the cell's current requirement for protein synthesis, but also the cellular stage of development.…”

“…Three types of oocyte maturation can be considered; cytoplasmic, nuclear, and epigenetic [ 11 ], all of which are essential for oocytes to acquire developmental competence. Oocyte epigenetic maturation is required for establishment of maternal imprinting [ 2 , 12 ], regulation of transcription [ 4 , 13 ], chromosome segregation [ 14 , 15 ] and to support EGA [ 9 , 12 , 16 ]. These are processes which take place at different times during oocyte and early embryo development.…”

Chromatin profiling and state predictions reveal insights into epigenetic regulation during early porcine development

“…Our analysis of H3K4me3 enrichment in PFF and PTr2 cells revealed that the vast majority of H3K4me3 sites in both cell lines were in promoter regions around transcriptional start sites of transcriptionally active genes, an observation that is consistent with the character of this mark as reported in the literature. H3K4me3 is one of the most widely profiled histone marks and is known to be enriched around active promoters during embryonic development and beyond [ 52 – 57 ]. Similarly, H3K27ac showed enrichment at promoter regions, though relatively less than H3K4me3, likely due to its presence at both promoters and enhancers [ 58 , 59 ].…”