The smallest repeating unit in the chromatin structure is a nucleosome. It consists of DNA which is wrapped in helical turns around a histone octamer. Single nucleosomes are connected via linker DNA, forming the next higher structure of chromatin, the so-called beads-on-a-string. Chromatin fulfills two tasks in the nucleus: packing 1.7 metres of DNA and limiting central nuclear DNA processes such as DNA replication, recombination, repair and transcription. The dynamic rearrangements of the chromatin are caused by chromatin remodelling complexes, which use the energy of ATP hydrolysis to alter chromatin structure.
We are interested in the packed as well as the dynamic structure of the chromatin which we investigate with a combination of single-molecule FRET and super-resolution microscopy.
→ Acetylation of histone H3 at lysine 64 regulates nucleosome dynamics and facilitates transcription
Post-translational modifications of proteins have emerged as a major mechanism for regulating gene expression. However, our understanding of how histone modifications directly affect chromatin function remains limited. In this study, we investigate acetylation of histone H3 at lysine 64 (H3K64ac), a previously uncharacterized acetylation on the lateral surface of the histone octamer. We show that H3K64ac regulates nucleosome stability and facilitates nucleosome eviction and hence gene expression in vivo. In line with this, we demonstrate that H3K64ac is enriched in vivo at the transcriptional start sites of active genes and it defines transcriptionally active chromatin. Moreover, we find that the p300 co-activator acetylates H3K64, and consistent with a transcriptional activation function, H3K64ac opposes its repressive counterpart H3K64me3. Our findings reveal an important role for a histone modification within the nucleosome core as a regulator of chromatin function and they demonstrate that lateral surface modifications can define functionally opposing chromatin states.
→ H2A.Z.2.2 is an alternatively spliced histone H2A.Z variant that causes severe nucleosome destabilization
The histone variant H2A.Z has been implicated in many biological processes, such as gene regulation and genome stability. Here, we present the identification of H2A.Z.2.2 (Z.2.2), a novel alternatively spliced variant of histone H2A.Z and provide a comprehensive
characterization of its expression and chromatin incorporation properties. Z.2.2 mRNA is found in all human cell lines and tissues with highest levels in brain. We show the proper splicing and in vivo existence of this variant protein in humans. Furthermore, we demonstrate the binding of Z.2.2 to H2A.Z-specific TIP60 and SRCAP chaperone complexes and its active replication-independent deposition into chromatin. Strikingly, various independent in vivo and in vitro analyses, such as biochemical fractionation, comparative FRAP studies of GFP-tagged H2A variants, size exclusion chromatography and single molecule FRET, in combination with in silico molecular dynamics simulations, consistently demonstrate that Z.2.2 causes major structural changes and significantly destabilizes nucleosomes. Analyses of deletion mutants and chimeric proteins pinpoint this property to its unique C-terminus. Our findings enrich the list of known human variants by an unusual protein belonging to the H2A.Z family that leads to the least stable nucleosome known to date.