Enhancer of Zeste is a Polycomb Group protein essential for the establishment and maintenance of repression of homeotic and other genes. In the early embryo it is found in a complex that includes ESC and is recruited to Polycomb Response Elements. We show that this complex contains a methyltransferase activity that methylates lysine 9 and lysine 27 of histone H3, but the activity is lost when the E(Z) SET domain is mutated. The lysine 9 position is trimethylated and this mark is closely associated with Polycomb binding sites on polytene chromosomes but is also found in centric heterochromatin, chromosome 4, and telomeric sites. Histone H3 methylated in vitro by the E(Z)/ESC complex binds specifically to Polycomb protein.
Histone modifications play an important role in shaping chromatin structure. Here, we describe the use of an in vitro chromatin assembly system from Drosophila embryo extracts to investigate the dynamic changes of histone modifications subsequent to histone deposition. In accordance with what has been observed in vivo, we find a deacetylation of the initially diacetylated isoform of histone H4, which is dependent on chromatin assembly. Immediately after deposition of the histones onto DNA, H4 is monomethylated at K20, which is required for an efficient deacetylation of the H4 molecule. H4K20 methylation-dependent dl(3)MBT association with chromatin and the identification of a dl(3)MBT-dRPD3 complex suggest that a deacetylase is specifically recruited to the monomethylated substrate through interaction with dl(3)MBT. Our data demonstrate that histone modifications are added and removed during chromatin assembly in a highly regulated manner.All DNA in a eukaryotic cell is assembled into chromatin to fit it into the restricted nuclear space and to organize the genome (29, 56). As the DNA content of a cell doubles during S-phase of the cell cycle, the cell has to provide sufficient histone molecules to package the newly replicated DNA into chromatin. This is achieved mainly by a coupling of histone and DNA synthesis (37). The progression of the DNA replication machinery disrupts the nucleosome in front of the replication fork, which is then reassembled onto the newly synthesized DNA strands in a random manner (15,20). The remaining gaps are subsequently filled up with newly translated histone molecules, leading to a mosaic pattern of new and old histone octamers (2, 21). This process is assisted by the action of nuclear chaperones, which bind to the histones before deposition (14, 34). The newly deposited histones are more loosely associated with the DNA than the bulk histones and mature slowly into a more stable chromatin structure. Assembly is achieved via an ordered deposition of H3 and H4, followed by the binding of H2A/H2B dimers and finally the interaction of the linker histone H1 with the chromatin fiber (19)(20)(21)(22)49).Posttranslational modifications of histone molecules are generally considered to play an important role during the establishment and maintenance of chromatin structures. The combination of histone modifications has been proposed to constitute a "histone code" (23, 55), which is involved in the maintenance of epigenetic information. However, it is unclear how histone modifications are replicated during cellular division and DNA repair when newly translated histones are deposited onto the DNA.Modification marks can be stably maintained through multiple mitotic divisions (3,23,30,55). However, for most histone modifications, the molecular mechanisms that regulate this maintenance have so far remained elusive. A key aspect of this maintenance or reestablishment of modification patterns is the precise copying of histone modification patterns from the parental histone to the newly synthesized on...
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