Linker histones play important roles in the genomic organization of mammalian cells. Of the linker histone variants, H1.X shows the most dynamic behavior in the nucleus. Recent research has suggested that the linker histone variants H1.X and H1.0 have different chromosomal binding site preferences. However, it remains unclear how the dynamics and binding site preferences of linker histones are determined. Here, we biochemically demonstrated that the DNA/nucleosome and histone chaperone binding activities of H1.X are significantly lower than those of other linker histones. This explains why H1.X moves more rapidly than other linker histones in vivo. Domain swapping between H1.0 and H1.X suggests that the globular domain (GD) and C-terminal domain (CTD) of H1.X independently contribute to the dynamic behavior of H1.X. Our results also suggest that the N-terminal domain (NTD), GD, and CTD cooperatively determine the preferential binding sites, and the contribution of each domain for this determination is different depending on the target genes. We also found that linker histones accumulate in the nucleoli when the nucleosome binding activities of the GDs are weak. Our results contribute to understanding the molecular mechanisms of dynamic behaviors, binding site selection, and localization of linker histones.
Histones, including linker histones, are highly basic proteins that randomly bind to DNA. To maintain genomic integrity, DNA binding of histones should be strictly controlled. It has been established that random DNA binding of histones abrogates the sequential nucleosome assembly process (1). Histone chaperones directly bind to histones and transfer them to DNA to assemble the chromatin structure. In addition, histone chaperones have been suggested to play an important role in inhibiting the unfavorable DNA binding of histones (1). Our previous studies identified template activating factor I (TAF-I), nucleosome assembly protein 1 (NAP1), and nucleophosmin/B23 as factors involved in regulating the transcription and replication of adenovirus chromatin (2, 3). Recently, all three proteins, TAF-I, NAP1, and B23, were found to function as linker histone chaperones (4-7). A nucleolar protein, nucleolin (NCL), and prothymosin ␣ (ProT␣) were shown to regulate linker histone-chromatin binding (8-10).Linker histones are comprised of three regions: the N-terminal domain (NTD), central globular domain (GD), and C-terminal domain (CTD). The GDs of about 80 amino acids are able to reside on the DNA entry and exit sites of a nucleosome (11). Recent structural analysis of the GD of chicken linker histone H5 demonstrated that the GD makes contact with two extended DNA strands from the nucleosome and the nucleosome dyad (12). The CTD of linker histones are generally basic and rich in lysines, alanines, and prolines. The CTD plays a critical role in the condensation of DNA through its random DNA binding activity (13,14). The GD and CTD show distinct DNA binding activity (15), but neither is sufficient to stably associate with nu...