A Phylogenetic Study of SPBP and RAI1: Evolutionary Conservation of Chromatin Binding Modules

Our genome is assembled into and array of highly dynamic nucleosome structures allowing spatial and temporal access to DNA. The nucleosomes are subject to a wide array of post-translational modifications, altering the DNA-histone interaction and serving as docking sites for proteins exhibiting effector or “reader” modules. The nuclear proteins SPBP and RAI1 are composed of several putative “reader” modules which may have ability to recognise a set of histone modification marks. Here we have performed a phylogenetic study of their putative reader modules, the C-terminal ePHD/ADD like domain, a novel nucleosome binding region and an AT-hook motif. Interactions studies in vitro and in yeast cells suggested that despite the extraordinary long loop region in their ePHD/ADD-like chromatin binding domains, the C-terminal region of both proteins seem to adopt a cross-braced topology of zinc finger interactions similar to other structurally determined ePHD/ADD structures. Both their ePHD/ADD-like domain and their novel nucleosome binding domain are highly conserved in vertebrate evolution, and construction of a phylogenetic tree displayed two well supported clusters representing SPBP and RAI1, respectively. Their genome and domain organisation suggest that SPBP and RAI1 have occurred from a gene duplication event. The phylogenetic tree suggests that this duplication has happened early in vertebrate evolution, since only one gene was identified in insects and lancelet. Finally, experimental data confirm that the conserved novel nucleosome binding region of RAI1 has the ability to bind the nucleosome core and histones. However, an adjacent conserved AT-hook motif as identified in SPBP is not present in RAI1, and deletion of the novel nucleosome binding region of RAI1 did not significantly affect its nuclear localisation.     

Determinants of histone H1 mobility and chromatin binding in living cells

The dynamic interaction of chromatin-binding proteins with their nucleosome binding sites is an important element in regulating the structure and function of chromatin in living cells. Here we review the major factors regulating the intranuclear mobility and chromatin binding of the linker histone H1, the most abundant family of nucleosome-binding proteins. The information available reveals that multiple and diverse factors modulate the interaction of H1 with chromatin at both a local and global level. This multifaceted mode of modulating the interaction of H1 with nucleosomes is part of the mechanism that regulates the dynamics of the chromatin fiber in living cells.     

Distinct domains in high mobility group N variants modulate specific chromatin modifications

We have demonstrated that levels of specific modification in histone H3 are modulated by members of the nucleosome-binding high mobility group N (HMGN) protein family in a variant-specific manner. HMGN1 (but not HMGN2) inhibits the phosphorylation of both H3S10 and H3S28, whereas HMGN2 enhances H3K14 acetylation more robustly than HMGN1. Two HMGN domains are necessary for modulating chromatin modifications, a non-modification-specific domain necessary for chromatin binding and a modification-specific domain localized in the C terminus of the HMGNs. Thus, chromatin-binding structural proteins such as HMGNs affect the levels of specific chromatin modifications and therefore may play a role in epigenetic regulation.     

High Mobility Group Protein N5 (HMGN5) and Lamina-associated Polypeptide 2α (LAP2α) Interact and Reciprocally Affect Their Genome-wide Chromatin Organization

The interactions of nuclear lamins with the chromatin fiber play an important role in regulating nuclear architecture and chromatin function; however, the full spectrum of these interactions is not known. We report that the N-terminal domain of the nucleosome-binding protein HMGN5 interacts with the C-terminal domain of the lamin-binding protein LAP2α and that these proteins reciprocally alter their interaction with chromatin. Chromatin immunoprecipitation analysis of cells lacking either HMGN5 or LAP2α reveals that loss of either protein affects the genome-wide distribution of the remaining partner. Our study identifies a new functional link between chromatin-binding and lamin-binding proteins.