Fundamental Role Of The H2A.Z C-Terminal Tail In The Formation Of Constitutive Heterochromatin

Nucleosome stability, a crucial determinant of gene regulation, was measured in a robust in situ assay to assess the molecular determinants of the stability of H2A.Z-containig nucleosomes. Surprisingly, a large fraction of H2A.Z detected by three different antibodies was released from the nucleosomes by salt together with H3, and was associated with H3K9me3 but not with H3K27me3 marked nucleosomes. This unusual behavior relied on the presence of the unstructured C-terminal chain of the histone variant, rather than on isoform specificity, posttranslational modifications or binding of the reader protein PWWPA2, as determined using cell lines expressing only particular forms of the variant. In the absence of this tail, or upon addition of an excess of the tail peptide to the nuclei of control cells, the canonical H2A-like stability features were readily restored and most of the H2A.Z-containing nucleosomes left the periphery and ended up in scattered foci in the nuclei. Concomitantly, the H3K9me3-marked constitutive heterochromatin was also dispersed, what was accompanied by increased overall nuclease sensitivity and significantly enhanced binding of intercalating dyes to the DNA. The DT40 cells expressing the tailless H2A.Z showed marked differences in their gene expression pattern and were distinguished by compromised DNA damage response. Thus, interactions involving a short H2A.Z peptide chain simultaneously determine the stability and accessibility features of chromatin involving the nucleosomes containing this histone variant and the localization of these large chromatin regions in the nucleus. Our data suggest that H2A.Z can function in both heterochromatic and in euchromatic scenarios depending on the molecular interactions involving its C-terminal unstructured tail, shedding light on the enigmatic double-faced character of this histone variant. ### Competing Interest Statement The authors have declared no competing interest.