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

Abstract

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.

The characterization of macroH2A beyond vertebrates supports an ancestral origin and conserved role for histone variants in chromatin

“…Z.2), another monophyletic lineage within H2A. 39,50,51 In both cases, the differentiation of specialized vertebrate variants is concomitant with an increase in organismal complexity, underscoring the role of these variants in refining chromatin structure and the associated epigenetic mechanisms. On the contrary, the apparent absence of macroH2A in some protostome groups (i.e., nematodes and holometabolous insects) indicates that this variant might not be indispensable for the correct development of all organisms.…”

Histone H2A variants in nucleosomes and chromatin: more or less stable?

“…In contrast, most histone variant mRNAs are polyadenylated, and their pre-mRNAs can contain introns (18). To date, only two histone transcripts have been shown to be alternatively spliced, macroH2A.1 (19) and H2A.Z.2 (20,21), giving rise to histone proteins with distinct functional and structural properties.…”

Chromatibody, a novel non-invasive molecular tool to explore and manipulate chromatin in living cells

“…Thus, the expression of a fluorescently-tagged histone has been a cuttingedge technique to visualize chromatin in cultured cells (Kanda et al, 1998), as well as in model organisms (Clarkson and Saint, 1999;Pauls et al, 2001) and still remains a common approach to study chromosome dynamics, mitosis or even apoptosis during developmental processes. Fluorescent-tagged histones that incorporate into nucleosomes have also been used to study their exchange dynamics within chromatin fiber (Kimura and Cook, 2001;Bonisch et al, 2012).…”