Live-cell analysis of DNA methylation during sexual reproduction in Arabidopsis reveals context and sex-specific dynamics controlled by noncanonical RdDM

Abstract

Cytosine methylation is a key epigenetic mark in many organisms, important for both transcriptional control and genome integrity. While relatively stable during somatic growth, DNA methylation is reprogrammed genome-wide during mammalian reproduction. Reprogramming is essential for zygotic totipotency and to prevent transgenerational inheritance of epimutations. However, the extent of DNA methylation reprogramming in plants remains unclear. Here, we developed sensors reporting with single-cell resolution CG and non-CG methylation in Arabidopsis. Live imaging during reproduction revealed distinct and sex-specific dynamics for both contexts. We found that CHH methylation in the egg cell depends on DOMAINS REARRANGED METHYLASE 2 (DRM2) and RNA polymerase V (Pol V), two main actors of RNA-directed DNA methylation, but does not depend on Pol IV. Our sensors provide insight into global DNA methylation dynamics at the single-cell level with high temporal resolution and offer a powerful tool to track CG and non-CG methylation both during development and in response to environmental cues in all organisms with methylated DNA, as we illustrate in mouse embryonic stem cells.

Epigenetic dynamics during flowering plant reproduction: evidence for reprogramming?

“…The targets of these processes are unknown and the epigenetic state of the MMC has not been profiled at molecular resolution. Recently, fusions between DNA methylation‐binding proteins and fluorescent reporters allowed visualization of methylation dynamics at a broad scale (Ingouff et al ., ). CG methylation remains steady but CHH methylation decreases transiently and is then restored during MMC specification.…”

“…Tansley insight New Phytologist fluorescent reporters allowed visualization of methylation dynamics at a broad scale (Ingouff et al, 2017). CG methylation remains steady but CHH methylation decreases transiently and is then restored during MMC specification.…”

“…As part of a recurring theme, AGO5 acts in a cell nonautonomous manner to promote the mitotic division of the functional megaspore (Tucker et al, 2012). Fluorescent reporters suggest that CG and CHH methylation remain fairly steady throughout female gametogenesis (Ingouff et al, 2017). The mature female gametophyte contains two gametes, a haploid egg cell and a diploid central cell.…”

Visualizing protein-protein interactions in plants by rapamycin-dependent delocalization

“…All primers sequences are shown in Supplemental Table 1 (Ingouff et al, 2017) and pUBQ (Mehlmer et al, 2012) promoters respectively. The eGFP was cloned into the pK7WG2 (Karimi et al, 2002(Karimi et al, , 2005.…”

SDG2-Mediated H3K4me3 Is Crucial for Chromatin Condensation and Mitotic Division during Male Gametogenesis in Arabidopsis

“…In mammals, an erasure of almost all DNA methylation states is observed before fertilization (Feng et al, 2010). In plants, however, such an extreme reprogramming event does not seem to occur at fertilization (Ingouff et al, 2017;Saze et al, 2003;Jullien et al, 2012;Calarco et al, 2012). Moreover, over 85% of mammalian histones are replaced by sperm-specific protamine proteins, enabling extensive compaction of DNA in male gametes, and upon fertilization the paternal chromatin is deprived of the protamines and loaded with variant histone H3.3 provided by the female gametes (Ooi and Henikoff, 2007;Saitou and Kurimoto, 2014).…”