Abstract
Acetylation of histone proteins is a major determinant of chromatin structure and function. Fertilisation triggers a round of chromatin remodelling that prepares the genome for the first round of transcription from the new embryonic genome. In this study we confirm that fertilisation leads to a marked progressive increase in the level of histone 3 lysine 9 acetylation in both the paternally and maternally derived genomes. The culture of zygotes in simple defined media caused a marked increase in the global level of acetylation and this affected the male pronucleus more than the female. The culture created a marked asymmetry in staining between the two pronuclei that was not readily detected in zygotes collected directly from the reproductive tract and was ameliorated to some extent by optimized culture media. The increased acetylation caused by culture resulted in increased transcription of Hspa1b, a marker of embryonic genome activation. Pharmacological analyses showed the hyperacetylation of H3K9 and the increased expression of Hspa1b caused by culture were due to the altered net activity of a range of histone acetylases and deacetylases. The marked hyperacetylation of histone 3 lysine 9 caused by culture of zygotes may serve as an early biomarker for the effects of culture on the normal function of the embryo. The results also provide further evidence for an effect of the stresses associated with assisted reproductive technologies on the normal patterns of epigenetic reprogramming in the early embryo.
Highlights
Activation of gene expression from the new embryonic genome created at fertilisation is an essential requirement for normal development
The results show that the onset of transcription of a marker of embryonic genome activation is in part regulated by rising level of global histone acetylation, and this process is perturbed by the culture of embryos in vitro
Confocal projections (Fig. 1A) of zygotes collected directly from the oviduct showed that H3K9ac staining was not detected in PN1 stage zygotes and was at a very low level at the PN2 stage in both pronuclei
Summary
Activation of gene expression from the new embryonic genome created at fertilisation is an essential requirement for normal development. This is a progressive process that, in the mouse, commences towards the end of the first cell cycle, and reaches maturity by the 8-cell stage (Latham & Schultz 2001). The new transcriptome created by this process has the essential roles of supporting the normal cellular functions present in all cells and has the unique feature of converting the transcribed genome of the terminally differentiated gametes into the totipotent state of the early embryo. It is generally considered that conversion to totipotency requires a process of extensive epigenetic reprogramming of the new embryonic genomes inherited from the gametes, there is currently a rather limited understanding of the essential details of this process. The efficiency of reprogramming after cloning is commonly quiet low, but treatment with a histone
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