Chromatin remodeling of histone H3 variants by DDM1 underlies epigenetic inheritance of DNA methylation.

Nucleosomes block access to DNA methyltransferase, unless they are remodeled by DECREASE in DNA METHYLATION 1 (DDM1), a Snf2-like master regulator of epigenetic inheritance. We show that DDM1 promotes replacement of histone variant H3.3 by H3.

Chromatin remodeling of histone H3 variants underlies epigenetic inheritance of DNA methylation.

Epigenetic inheritance refers to the faithful replication of DNA methylation and histone modification independent of DNA sequence. Nucleosomes block access to DNA methyltransferases, unless they are remodeled by DECREASE IN DNA METHYLATION1 (DDM1 ), a Snf2-like master regulator of epigenetic inheritance. We show that DDM1 activity results in replacement of the transcriptional histone variant H3.

Nucleosome Turnover Regulates Histone Methylation Patterns over the Genome.

Recent studies have indicated that nucleosome turnover is rapid, occurring several times per cell cycle. To access the effect of nucleosome turnover on the epigenetic landscape, we investigated H3K79 methylation, which is produced by a single methyltransferase (Dot1l) with no known demethylase. Using chemical-induced proximity (CIP), we find that the valency of H3K79 methylation (mono-, di-, and tri-) is determined by nucleosome turnover rates.

Rapid and reversible epigenome editing by endogenous chromatin regulators.

Understanding the causal link between epigenetic marks and gene regulation remains a central question in chromatin biology. To edit the epigenome we developed the FIRE-Cas9 system for rapid and reversible recruitment of endogenous chromatin regulators to specific genomic loci. We enhanced the dCas9-MS2 anchor for genome targeting with Fkbp/Frb dimerizing fusion proteins to allow chemical-induced proximity of a desired chromatin regulator.

TOP2 synergizes with BAF chromatin remodeling for both resolution and formation of facultative heterochromatin.

The resolution and formation of facultative heterochromatin are essential for development, reprogramming, and oncogenesis. The mechanisms underlying these changes are poorly understood owing to the difficulty of studying heterochromatin dynamics and structure in vivo. We devised an in vivo approach to investigate these mechanisms and found that topoisomerase II (TOP2), but not TOP1, synergizes with BAF (mSWI/SNF) ATP-dependent chromatin remodeling complexes genome-wide to resolve facultative heterochromatin to accessible chromatin independent of transcription.

Dynamics of BAF-Polycomb complex opposition on heterochromatin in normal and oncogenic states.

The opposition between Polycomb repressive complexes (PRCs) and BAF (mSWI/SNF) complexes has a critical role in both development and disease. Mutations in the genes encoding BAF subunits contribute to more than 20% of human malignancies, yet the underlying mechanisms remain unclear, owing largely to a lack of assays to assess BAF function in living cells. To address this, we have developed a widely applicable recruitment assay system through which we find that BAF opposes PRC by rapid, ATP-dependent eviction, leading to the formation of accessible chromatin.

Smarca4 ATPase mutations disrupt direct eviction of PRC1 from chromatin.

Trithorax-group proteins and their mammalian homologs, including those in BAF (mSWI/SNF) complexes, are known to oppose the activity of Polycomb repressive complexes (PRCs). This opposition underlies the tumor-suppressive role of BAF subunits and is expected to contribute to neurodevelopmental disorders. However, the mechanisms underlying opposition to Polycomb silencing are poorly understood.

RNA-directed DNA methylation regulates parental genomic imprinting at several loci in Arabidopsis.

In mammals and plants, parental genomic imprinting restricts the expression of specific loci to one parental allele. Imprinting in mammals relies on sex-dependent de novo deposition of DNA methylation during gametogenesis but a comparable mechanism was not shown in plants. Rather, paternal silencing by the maintenance DNA methyltransferase 1 (MET1) and maternal activation by the DNA demethylase DEMETER (DME) cause maternal expression.

Imprinting: DNA methyltransferases illuminate reprogramming.

Progress in studying epigenetic reprogramming in plants has been impeded by the difficulty in obtaining tissue for analysis. Now, using a combination of fluorescent reporters and translational fusions, a new study sheds some light on this process.

FACS-based purification of Arabidopsis microspores, sperm cells and vegetative nuclei.

Background: The male germline in flowering plants differentiates by asymmetric division of haploid uninucleated microspores, giving rise to a vegetative cell enclosing a smaller generative cell, which eventually undergoes a second mitosis to originate two sperm cells. The vegetative cell and the sperm cells activate distinct genetic and epigenetic mechanisms to control pollen tube growth and germ cell specification, respectively. Therefore, a comprehensive characterization of these processes relies on efficient methods to isolate each of the different cell types throughout male gametogenesis.

Reprogramming of DNA methylation in pollen guides epigenetic inheritance via small RNA.

Epigenetic inheritance is more widespread in plants than in mammals, in part because mammals erase epigenetic information by germline reprogramming. We sequenced the methylome of three haploid cell types from developing pollen: the sperm cell, the vegetative cell, and their precursor, the postmeiotic microspore, and found that unlike in mammals the plant germline retains CG and CHG DNA methylation. However, CHH methylation is lost from retrotransposons in microspores and sperm cells and restored by de novo DNA methyltransferase guided by 24 nt small interfering RNA, both in the vegetative nucleus and in the embryo after fertilization.

Genome reprogramming and small interfering RNA in the Arabidopsis germline.

The movement of mobile small RNA signals between cells has garnered much interest over the last few years, and has recently been extended to germ cells during gamete development. Focusing on plants, we review mobile RNA signals that arise following reprogramming in the germline, and their effect on transposable element silencing on the one hand and on meiotic and apomictic germ cell fate on the other. A potential role for reprogramming and small RNA in hybrid formation and speciation is proposed.

Global analysis of alternative splicing differences between humans and chimpanzees.

Alternative splicing is a powerful mechanism affording extensive proteomic and regulatory diversity from a limited repertoire of genes. However, the extent to which alternative splicing has contributed to the evolution of primate species-specific characteristics has not been assessed previously. Using comparative genomics and quantitative microarray profiling, we performed the first global analysis of alternative splicing differences between humans and chimpanzees.

Par6-aPKC uncouples ErbB2 induced disruption of polarized epithelial organization from proliferation control.

The polarized glandular organization of epithelial cells is frequently lost during development of carcinoma. However, the specific oncogene targets responsible for polarity disruption have not been identified. Here, we demonstrate that activation of ErbB2 disrupts apical-basal polarity by associating with Par6-aPKC, components of the Par polarity complex.

Reversible photocontrol of DNA binding by a designed GCN4-bZIP protein.

Synthetic photocontrolled proteins could be powerful tools for probing cellular chemistry. Several previous attempts to produce such systems by incorporating photoisomerizable chromophores into biomolecules have led to photocontrol but with incomplete reversibility, where the chromophore becomes trapped in one photoisomeric state. We report here the design of a modified GCN4-bZIP DNA-binding protein with an azobenzene chromophore introduced between Cys residues at positions 262 and 269 (S262C, N269C) within the zipper domain.