Polycomb group proteins

Epigenetic regulation of chromatin structure is essential for the expression of genes determining cellular specification and function. The Polycomb repressive complex 2 (PRC2) di- and trimethylates histone H3 on lysine 27 (H3K27me2/me3) to establish repression of specific genes in embryonic stem cells and during differentiation. How the Polycomb group (PcG) target genes are regulated by environmental cues and signaling pathways is quite unexplored. Here, we show that the mitogen- and stress-activated kinases (MSK), through a mechanism that involves promoter recruitment, histone H3K27me3S28 phosphorylation, and displacement of PcG proteins, lead to gene activation. We present evidence that the H3K27me3S28 phosphorylation is functioning in response to stress signaling, mitogenic signaling, and retinoic acid (RA)-induced neuronal differentiation. We propose that MSK-mediated H3K27me3S28 phosphorylation serves as a mechanism to activate a subset of PcG target genes determined by the biological stimuli and thereby modulate the gene expression program determining cell fate.► The H3K27me3S28p double mark exists in vivo ► Mitogen and stress-activated kinases phosphorylates H3K27me3S28 to displace PcG proteins ► H3K27me3S28 phosphorylation is induced in response to mitogen, stress, and differentiation signals ► H3K27me3S28 phosphorylation correlates with transcriptional activation of target genes

Polycomb group (PcG) proteins are part of a transcriptional memory system in
higher eukaryotes, and are necessary to maintain the transcriptional repressed states
of HOX genes. PcG proteins also have a well-known effect on the cell-cycle via
repression of the Cdkn2a/INK4A locus. In order to repress genes the EED/EZH2 PcG
complex deactelyates the histone H3 N-terminal tail, and induces trimethylation of
histone H3 at lysine 27 (H3K27
3M), known to be bound by the human Polycomb
Repressive Complex 1 (hPRC1). Little is known about the regulation of PcG proteins
themselves. PcG proteins are regulated both cell-cycle dependently and
independently. Cell-cycle dependent phosphorylation of the PcG protein Bmi1
correlates inversely with its chromatin association.53 Cell-cycle independent
regulation involves the mitogen activated protein kinase (MAPK) cascades, all
inducing a MAPK activated protein kinase called 3pK, which is an in vivo kinase for
Bmi1.
In the present study we found that stress signaling results in loss of
pericentromeric association of the PcG proteins Bmi1, Ring1A, Ring1B and HPc2.
The PcG protein HPH1 shows a nuclear redistribution upon activation of MAPK
signaling. MAPK induced (cell-cycle independent) phosphorylation of the PcG protein
Bmi1 in addition was shown to result in chromatin dissociation, while HPH1 only
detaches in part form chromatin under these conditions. Immunofluorescence data
indicate that MAPK induced phsophorylation of H3 at serine 28 might be involved in
chromatin dissociation of PcG proteins from the H3K27
3M bound hPRC1 complex.
Upon chromatin dissociation interactions with 3pK are maintained intact.
Preliminary data show that ubiquitination might play a role in the PcG
transcriptional memory system. Overexpression of TAP (Tandem Affinity Purification)
tagged Ring1B (an in vitro ubiquitin E3 ligase) resulted in a 7 kDa increase by 3pK
immunodetection on a Western blot, also detected with an ubiquitin specific antibody,
and pulled down with antisera against 3pK. Further experiments are necessary to
verify whether Ring1B is involved in mono-ubiquitination of 3pK, and to test whether
the hPRC1 complex is involved in histone ubiquitination.

The Drosophila spermatogenesis cell differentiation pathway involves the activation of a large set of genes in primary spermatocytes. Most of these genes are activated by testis-specific TATA-binding protein associated factors (tTAFs). In the current model for the activation mechanism, Polycomb plays a key role silencing these genes in the germline precursors, and tTAF-dependent activation in primary spermatocytes involves the displacement of Polycomb from gene promoters. We investigated the genome-wide binding of Polycomb in wild type and tTAF mutant testes. According to the model we expected to see a clear enhancement in Polycomb binding at tTAF-dependent spermatogenesis genes in tTAF mutant testes. However, we find little evidence for such an enhancement in tTAF mutant testes compared to wild type. To avoid problems arising from cellular heterogeneity in whole testis analysis, we further tested the model by analysing Polycomb binding in purified germline precursors, representing ...

Polycomb group (PcG) proteins are key regulators in establishing a transcriptional repressive state. Polycomb
Repressive Complex 2 (PRC2), one of the two major PcG protein complexes, is essential for proper
differentiation and maintenance of cellular identity. Multiple factors are involved in recruiting PRC2
to its genomic targets. In this review, we will discuss the role of DNA sequence, transcription factors,
pre-existing histone modifications, and RNA in guiding PRC2 towards specific genomic loci. The DNA
sequence itself influences the DNA methylation state, which is an important determinant of PRC2 recruitment.
Other histone modifications are also important for PRC2 binding as PRC2 can respond to different
cellular states via crosstalk between histone modifications. Additionally, PRC2 might be able to sense
the transcriptional status of genes by binding to nascent RNA, which could also guide the complex to
chromatin. In this review, we will discuss how all these molecular aspects define a local chromatin state
which controls accurate, cell-type-specific epigenetic silencing by PRC2.

The RING domain protein Arkadia/RNF111 is a ubiquitin ligase in the TGFβ pathway. We previously identified Arkadia as a SUMO-binding protein with clustered SUMO-interacting motifs (SIMs) that together form a SUMO-binding domain (SBD). However, precisely how SUMO interaction contributes to the function of Arkadia was not resolved. Through analytical molecular and cell biology, we found that the SIMs share redundant function with Arkadia M-domain, a region distinguishing Arkadia from its paralogs ARKL1/ARKL2 and the prototypical SUMO-targeted ubiquitin ligase (STUbL) RNF4. The SIMs and M-domain together promote both Arkadia's colocalization with CBX4/Pc2, a component of Polycomb bodies, and the activation of a TGFβ pathway transcription reporter. Transcriptome profiling through RNA-sequencing showed that Arkadia can both promote and inhibit gene expression, indicating that Arkadia's activity in transcriptional control may depend on epigenetic context defined by Polycomb repres...

The Polycomb group (Pc-G) constitutes an important, functionally conserved group of proteins, required to stably maintain inactive homeobox genes repressed during development. Drosophila extra sex combs (esc) and its mammalian homolog embryonic ectoderm development (eed) are special Pc-G members, in that they are required early during development when Pc-G repression is initiated, a process that is still poorly understood. To get insight in the molecular function of Eed, we searched for Eed-interacting proteins, using the yeast two-hybrid method. Here we describe the specific in vivo binding of Eed to Enx1 and Enx2, two mammalian homologs of the essential Drosophila Pc-G gene Enhancer-of-zeste [E(z)]. No direct biochemical interactions were found between Eed/Enx and a previously characterized mouse Pc-G protein complex, containing several mouse Pc-G proteins including mouse polyhomeotic (Mph1). This suggests that different Pc-G complexes with distinct functions may exist. However, p...