Higher chromatin mobility supports totipotency and precedes pluripotency in vivo

Abstract

The fusion of the gametes upon fertilization results in the formation of a totipotent cell. Embryonic chromatin is expected to be able to support a large degree of plasticity. However, whether this plasticity relies on a particular conformation of the embryonic chromatin is unknown. Moreover, whether chromatin plasticity is functionally linked to cellular potency has not been addressed. Here, we adapted fluorescence recovery after photobleaching (FRAP) in the developing mouse embryo and show that mobility of the core histones H2A, H3.1, and H3.2 is unusually high in two-cell stage embryos and decreases as development proceeds. The transition toward pluripotency is accompanied by a decrease in histone mobility, and, upon lineage allocation, pluripotent cells retain higher mobility than the differentiated trophectoderm. Importantly, totipotent two-cell-like embryonic stem cells also display high core histone mobility, implying that reprogramming toward totipotency entails changes in chromatin mobility. Our data suggest that changes in chromatin dynamics underlie the transitions in cellular plasticity and that higher chromatin mobility is at the nuclear foundations of totipotency.Supplemental material is available for this article.Received January 25, 2014; revised version accepted April 16, 2014. Embryonic cells are characterized by a large degree of plasticity, which is the ability to generate different cell types upon differentiation and is necessary to start a full developmental program. After fertilization, the mouse embryo has the transient capacity to generate both embryonic and extraembryonic cell types, a feature that is referred to as totipotency (Tarkowski 1959;Ishiuchi and Torres-Padilla 2013). This is in contrast to pluripotent cells, which contribute to all three germ layers of the embryo proper, but not to extraembryonic lineages, and therefore have a more restricted potential than totipotent cells. In mice, only the zygote and two-cell stage blastomeres are, strictly speaking, totipotent, since they have the ability to develop into a full organism without the need of carrier cells (Tarkowski 1959;Tarkowski and Wroblewska 1967;Kelly et al. 1978;Ishiuchi and Torres-Padilla 2013). As development progresses, pluripotent cells form in the inner cell mass (ICM) of the blastocyst, accompanied by the activation of pluripotency-associated transcription factors like Nanog and Pou5f1/Oct4 (Nichols et al. 1998;Chambers et al. 2003). The first differentiated embryonic tissue, the trophectoderm (TE), appears morphologically distinguishable and surrounds the ICM in the blastocyst. Thus, during the early stages of development, the mouse embryo undergoes dramatic changes in cellular plasticity.Upon fertilization, embryonic chromatin undergoes intense chromatin remodeling. Indeed, this epigenetic reprogramming of the gametes is thought to be essential to gain totipotency (Sado and Ferguson-Smith 2005;Surani et al. 2007;Hemberger et al. 2009). However, the precise conformation of embryonic chromatin and the way it...

The pluripotency factor Nanog regulates pericentromeric heterochromatin organization in mouse embryonic stem cells

“…Constitutive heterochromatin is rapidly compacted upon ESC differentiation and embryo development, implying close coordination of chromatin with cell status (Meshorer et al 2006;Ahmed et al 2010;Boskovic et al 2014). Our findings suggest that down-regulation of Nanog, one of the earliest events in ESC differentiation, is a key driver of heterochromatin compaction.…”

“…All qPCR data represent mean ± SD from three biological experiments. Boskovic et al 2014). We identified a critical new role for the transcription factor Nanog in the establishment and maintenance of open heterochromatin, thereby forming a direct link between the ESC regulatory network and nuclear organization in pluripotent cells.…”

“…Pluripotent mouse embryonic stem cell (ESC) chromatin exists in an unusual configuration with widely dispersed open chromatin throughout the nucleoplasm, including within constitutive heterochromatin domains such as pericentromeric satellite repeats (Meshorer et al 2006;Efroni et al 2008;Fussner et al 2011;de Wit et al 2013). A similar form of highly dispersed chromatin architecture also characterizes pluripotent epiblast cells within the mouse blastocyst (Ahmed et al 2010;Boskovic et al 2014). Upon cell differentiation, there is extensive nuclear reorganization that is associated with chromatin compaction and the formation of condensed heterochromatin domains that form a repressive environment (Meshorer et al 2006;Efroni et al 2008;Wen et al 2009;Ahmed et al 2010;Wijchers et al 2015).…”

Building up the nucleus: nuclear organization in the establishment of totipotency and pluripotency during mammalian development

“…Current knowledge from the preimplantation embryo regarding the dynamics of the nucleus is restricted to the core mobility of histones. Histone mobility is higher in the totipotent two-cell stage embryo compared with the more pluripotent eight-cell stage nuclei (Boskovic et al 2014), in line with a higher histone core mobility of ESCs compared with differentiated cells (Meshorer et al 2006). Additionally, lamin B1 shows a higher turnover in ESCs than in MEFs as measured by fluorescence recovery after photobleaching (FRAP), suggesting a higher plasticity of the nuclear lamina in pluripotent cells as compared with somatic cells (Bhattacharya et al 2009).…”

Histone H3 Dynamics Reveal Domains with Distinct Proliferation Potential in the Arabidopsis Root

“…It is remarkable that the same principle applies to the end of cell proliferation and the end of the endocycle program, which are very different processes (Edgar et al, 2014). Interestingly, distinct degrees of chromatin dynamics have been reported during reprogramming at very early stages of mouse development (Bo sković et al, 2014;Burton and TorresPadilla, 2014) and along the root (Rosa et al, 2014).…”