The DNA-binding polycomb group protein pleiohomeotic mediates silencing of a Drosophila homeotic gene.

Polycomb group (PcG) proteins repress homeotic genes in cells where these genes must remain inactive during development. This repression requires cis-acting silencers, also called PcG response elements. Currently, these silencers are ill-defined sequences and it is not known how PcG proteins associate with DNA. Here, we show that the Drosophila PcG protein Pleiohomeotic binds to specific sites in a silencer of the homeotic gene Ultrabithorax. In an Ultrabithorax reporter gene, point mutations in these Pleiohomeotic binding sites abolish PcG repression in vivo. Hence, DNA-bound Pleiohomeotic protein may function in the recruitment of other non-DNA-binding PcG proteins to homeotic gene silencers.     

Polycomb group and trithorax group proteins in Arabidopsis

Polycomb group (PcG) and trithorax group (trxG) proteins form molecular modules of a cellular memory mechanism that maintains gene expression states established by other regulators. In general, PcG proteins are responsible for maintaining a repressed expression state, whereas trxG proteins act in opposition to maintain an active expression state. This mechanism, first discovered in Drosophila and subsequently in mammals, has more recently been studied in plants. The characterization of several Polycomb Repressive Complex 2 (PRC2) components in Arabidopsis thaliana constituted a first breakthrough, revealing key roles of PcG proteins in the control of crucial plant developmental processes. Interestingly, the recent identification of plant homologues of the Drosophila trithorax protein suggests a conservation of both the PcG and trxG gene regulatory system in plants. Here, we review the current evidence for the role of PcG and trxG proteins in the control of plant development, their biochemical functions, their interplay in maintaining stable expression states of their target genes, and point out future directions which may help our understanding of PcG and trxG function in plants.     

Molecular and genetic analysis of the Polycomb group gene Sex combs extra/Ring in Drosophila

Polycomb group (PcG) proteins repress homeotic genes and other developmental regulatory genes in cells where these genes must remain inactive during development. In Drosophila and in vertebrates, PcG proteins exist in two distinct multiprotein complexes, the Esc/Eed-E(z) complex and PRC1. Drosophila PRC1 contains Polycomb, Posterior sexcombs and Polyhomeotic, the products of three PcG genes that are critically needed for PcG silencing. Formation of stable PRC1 requires Ring, the product of a gene for which no mutations have been described. Here, we show that Sex combs extra (Sce) encodes Ring and that Sce/Ring function is critically required for PcG silencing.     

Telomeric associated sequences of Drosophila recruit polycomb-group proteins in vivo and can induce pairing-sensitive repression

In Drosophila, relocation of a euchromatic gene near centromeric or telomeric heterochromatin often leads to its mosaic silencing. Nevertheless, modifiers of centromeric silencing do not affect telomeric silencing, suggesting that each location requires specific factors. Previous studies suggest that a subset of Polycomb-group (PcG) proteins could be responsible for telomeric silencing. Here, we present the effect on telomeric silencing of 50 mutant alleles of the PcG genes and of their counteracting trithorax-group genes. Several combinations of two mutated PcG genes impair telomeric silencing synergistically, revealing that some of these genes are required for telomeric silencing. In situ hybridization and immunostaining experiments on polytene chromosomes revealed a strict correlation between the presence of PcG proteins and that of heterochromatic telomeric associated sequences (TASs), suggesting that TASs and PcG complexes could be associated at telomeres. Furthermore, lines harboring a transgene containing an X-linked TAS subunit and the mini-white reporter gene can exhibit pairing-sensitive repression of the white gene in an orientation-dependent manner. Finally, an additional binding site for PcG proteins was detected at the insertion site of this type of transgene. Taken together, these results demonstrate that PcG proteins bind TASs in vivo and may be major players in Drosophila telomeric position effect (TPE).     

Reconstitution of a functional core polycomb repressive complex

The opposing actions of polycomb (PcG) and trithorax group (trxG) gene products maintain essential gene expression patterns during Drosophila development. PcG proteins are thought to establish repressive chromatin structures, but the mechanisms by which this occurs are not known. Polycomb repressive complex 1 (PRC1) contains several PcG proteins and inhibits chromatin remodeling by trxG-related SWI/SNF complexes. We have defined a functional core of PRC1 by reconstituting a stable complex using four recombinant PcG proteins. One subunit, PSC, can also inhibit chromatin remodeling on its own. These PcG proteins create a chromatin structure that has normal nucleosome organization and is accessible to nucleases but excludes hSWI/SNF.     

Polycomb-dependent regulatory contacts between distant Hox loci in Drosophila

In Drosophila melanogaster, Hox genes are organized in an anterior and a posterior cluster, called Antennapedia complex and bithorax complex, located on the same chromosome arm and separated by 10 Mb of DNA. Both clusters are repressed by Polycomb group (PcG) proteins. Here, we show that genes of the two Hox complexes can interact within nuclear PcG bodies in tissues where they are corepressed. This colocalization increases during development and depends on PcG proteins. Hox gene contacts are conserved in the distantly related Drosophila virilis species and they are part of a large gene interaction network that includes other PcG target genes. Importantly, mutations on one of the loci weaken silencing of genes in the other locus, resulting in the exacerbation of homeotic phenotypes in sensitized genetic backgrounds. Thus, the three-dimensional organization of Polycomb target genes in the cell nucleus stabilizes the maintenance of epigenetic gene silencing.     

Diverse functions of Polycomb group proteins during plant development

Polycomb group (PcG) proteins play essential roles in animal and plant life cycles by controlling the expression of important developmental regulators. These structurally heterogeneous proteins form multimeric protein complexes that control higher order chromatin structure and, thereby, the expression state of their target genes. Once established, PcG proteins maintain silent gene expression states over many cell divisions providing a molecular basis for a cellular 'memory.' PcG proteins are best known for their role in the control of homeotic genes in Drosophila and mammals. In addition, they play important roles in the control of cell proliferation in vertebrate and invertebrate systems. Recent studies in plants have shown that PcG proteins regulate diverse developmental processes and, as in animals, they affect both homeotic gene expression and cell proliferation. Thus, the function of PcG proteins has been widely conserved between the plant and animal kingdoms.     

Association of trxG and PcG proteins with the bxd maintenance element depends on transcriptional activity

Polycomb group (PcG) and trithorax group (trxG) proteins act in an epigenetic fashion to maintain active and repressive states of expression of the Hox and other target genes by altering their chromatin structure. Genetically, mutations in trxG and PcG genes can antagonize each other's function, whereas mutations of genes within each group have synergistic effects. Here, we show in Drosophila that multiple trxG and PcG proteins act through the same or juxtaposed sequences in the maintenance element (ME) of the homeotic gene Ultrabithorax. Surprisingly, trxG or PcG proteins, but not both, associate in vivo in any one cell in a salivary gland with the ME of an activated or repressed Ultrabithorax transgene, respectively. Among several trxG and PcG proteins, only Ash1 and Asx require Trithorax in order to bind to their target genes. Together, our data argue that at the single-cell level, association of repressors and activators correlates with gene silencing and activation, respectively. There is, however, no overall synergism or antagonism between and within the trxG and PcG proteins and, instead, only subsets of trxG proteins act synergistically.     

A novel human polycomb binding site acts as a functional polycomb response element in Drosophila

Polycomb group (PcG) proteins are key chromatin regulators implicated in multiple processes including embryonic development, tissue homeostasis, genomic imprinting, X-chromosome inactivation, and germ cell differentiation. The PcG proteins recognize target genomic loci through cis DNA sequences known as Polycomb Response Elements (PREs), which are well characterized in Drosophila. However, mammalian PREs have been elusive until two groups reported putative mammalian PREs recently. Consistent with the existence of mammalian PREs, here we report the identification and characterization of a potential PRE from human T cells. The putative human PRE has enriched binding of PcG proteins, and such binding is dependent on a key PcG component SUZ12. We demonstrate that the putative human PRE carries both genetic and molecular features of Drosophila PRE in transgenic flies, implying that not only the trans PcG proteins but also certain features of the cis PREs are conserved between mammals and Drosophila.     

Structure of a polycomb response element and in vitro binding of polycomb group complexes containing GAGA factor

Polycomb response elements (PREs) are regulatory sites that mediate the silencing of homeotic and other genes. The bxd PRE region from the Drosophila Ultrabithorax gene can be subdivided into subfragments of 100 to 200 bp that retain different degrees of PRE activity in vivo. In vitro, embryonic nuclear extracts form complexes containing Polycomb group (PcG) proteins with these fragments. PcG binding to some fragments is dependent on consensus sequences for the GAGA factor. Other fragments lack GAGA binding sites but can still bind PcG complexes in vitro. We show that the GAGA factor is a component of at least some types of PcG complexes and may participate in the assembly of PcG complexes at PREs.     

PRE-mediated bypass of two Su(Hw) insulators targets PcG proteins to a downstream promoter

Drosophila Polycomb group response elements (PRE) silence neighboring genes, but silencing can be blocked by one copy of the Su(Hw) insulator element. We show here that Polycomb group (PcG) proteins can spread from a PRE in the flanking chromatin region and that PRE blocking depends on a physical barrier established by the insulator to PcG protein spreading. On the other hand, PRE-mediated silencing can bypass two Su(Hw) insulators to repress a downstream reporter gene. Strikingly, insulator bypass involves targeting of PcG proteins to the downstream promoter, while they are completely excluded from the intervening insulated domain. This shows that PRE-dependent silencing is compatible with looping of the PRE in order to bring PcG proteins in contact with the promoter and does not require the coating of the whole chromatin domain between PRE and promoter.     

The Drosophila pleiohomeotic mutation enhances the Polycomblike and Polycomb mutant phenotypes during embryogenesis and in the adult

In Drosophila, the spatially restricted expression of the homeotic genes is controlled by Polycomb group (PcG) repression. PcG proteins appear to form different complexes to repress this gene expression. Although the pleiohomeotic gene (pho) shares mutational phenotypes with other PcG mutations, which demonstrates that PHO binds directly with a Polycomb (Pc)-containing complex, the genetic interactions of pho with other PcG genes have not been examined in detail. Here we investigated whether pho interacts with Polycomblike (Pcl) and Polycomb (Pc) during embryonic and adult development using developmental and genetic approaches. Pcl and Pc strongly enhanced pho phenotypes in the legs and tergite of the adult fly. Embryonic cuticle transformation was also greatly enhanced in Pcl; pho or Pc; pho double mutant embryos. The double mutant phenotypes were more severely affected by the pho maternal effect mutation than in zygotic mutant background, suggesting dosage-dependent processes. Taken together, these results provide genetic evidence of an interaction between PHO with other Polycomb group proteins at the embryonic and adult stages, and of the functioning of PHO as a component of the PcG complex.