H2A.Z.2.2 is an alternatively spliced H2A.Z variant that causes severe nucleosome destabilization

Department of Molecular Biology, Adolf-Butenandt-Institute, Ludwig-Maximilians-University Munich, 80336 Munich, Germany.
Nucleic Acids Research (Impact Factor: 9.11). 03/2012; 40(13):5951-64. DOI: 10.1093/nar/gks267
Source: PubMed


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.

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    • "Only bursts with more than 50 photons in total were considered for further analysis. The populations of singly-labeled molecules were excluded by a stoichiometry (Sto) threshold of 0.1<Sto<0.7 and rare multi-molecule events were excluded by limiting the time deviation signals (TDS) to TDS<1 and TDS red-PIE <0.4 as explained in Bönisch et al. (2012). Because of the chosen position of the dyes, intact nucleosomes showed an intermediate to high FRET efficiency, while disassembled nucleosomes showed a very low FRET efficiency (dyes position shown in Figure 5A). "
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    Full-text · Article · Mar 2014 · eLife Sciences
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