Histone H2A Ubiquitination Does Not Preclude Histone H1 Binding, but It Facilitates Its Association with the Nucleosome

Abstract

Histone H2A ubiquitination is a bulky posttranslational modification that occurs at the vicinity of the binding site for linker histones in the nucleosome. Therefore, we took several experimental approaches to investigate the role of ubiquitinated H2A (uH2A) in the binding of linker histones. Our results showed that uH2A was present in situ in histone H1-containing nucleosomes. Notably in vitro experiments using nucleosomes reconstituted onto 167-bp random sequence and 208-bp (5 S rRNA gene) DNA fragments showed that ubiquitination of H2A did not prevent binding of histone H1 but it rather enhanced the binding of this histone to the nucleosome. We also showed that ubiquitination of H2A did not affect the positioning of the histone octamer in the nucleosome in either the absence or the presence of linker histones.Despite the renewed interest in histone H2A/H2B ubiquitination (1-3), the functional role of uH2A 1 still remains controversial. This is in contrast to ubiquitinated H2B where a strong correlation with transcriptional activation has long been established (2), and even a molecular mechanism involving a transhistone regulatory pathway has been shown to be involved in this process (4, 5). With uH2A, there is almost as much experimental evidence for its association to actively transcribing chromatin as there is to the opposite. For instance, it was shown that nucleosomes from the transcriptionally poised hsp70 and copia genes from Drosophila contain 50% uH2A (6). Also nucleosomes containing mono-and diubiquitinated H2A were found to preferentially occur near the 5Ј-end of the transcribed mouse dihydrofolate reductase gene (7). On the other hand, early studies with rat liver nuclei linked the disappearance of uH2A to increased transcription (8), and it was shown that Drosophila nucleosomes consisting of non-transcribed 1.705 satellite DNA showed an enrichment in uH2A (9). In support of these early data, a role for histone uH2A in Polycomb silencing has been demonstrated recently (10). Furthermore uH2A has been shown to be concentrated in the heterochromatic inactive sex body in pachytene spermatocytes (11). Nevertheless the molecular mechanism(s) involved remains completely unknown.Despite the controversy, ubiquitinated histones have been associated with chromatin partially depleted of linker histones (12) and transcriptionally active or poised sequences (1, 2, 6, 7, 13) that are thought to be have a reduced linker histone content or have altered association with linker histones (for a review, see Ref. 14). However, there is evidence that suggests that histone H2A ubiquitination does not interfere with linker histone binding. Two independent studies have shown that H1 can be cross-linked to uH2A in vitro (15) and in vivo (16). Moreover, in the cross-linking studies performed in mouse cells, the molar ratio of H1-uH2A to H1-H2A was the same as the molar ratio of uH2A to H2A (16).Linker histones are required for the stabilization of well defined chromatin fibers (17-19), and interference with the binding of lin...

Ubiquitination of histone H2B regulates chromatin dynamics by enhancing nucleosome stability

“…For instance, H2Aub1 facilitates the binding of linker histone H1 to reconstituted nucleosomes (33); and its absence releases H1 from chromatin (34). Collectively, all these findings support the view that histone ubiquitination plays an important role in controlling chromatin structure by regulating nucleosome stability.…”

Nucleosome landscape and control of transcription in the human malaria parasite

“…All together these data indicate that post-transcriptional mechanisms may be major mechanisms of gene regulation in the asexual development of the parasite. The classical transcriptiondriven eukaryotic model of gene regulation was already challenged by the relative paucity of transcription factors (Coulson et al 2004) and cis-acting regulatory elements detected in the parasite genome (Jason et al 2005;De Silva et al 2008). Moreover, even though our chromatin dynamic analyses at the binding sites of the ApiAP2 transcription factors support a role for transcriptional regulation (De Silva et al 2008), our results indicate that these transcription factors are following the general pattern found for most genes (cluster I).…”

Role of polycomb proteins Ring1A and Ring1Bin the epigenetic regulation of gene expression

“…It would be interesting to investigate whether the repression of the bivalent domains identified in differentiated cells (Azuara, 2006;Barski, 2007;Mikkelsen, 2007;Pan, 2007;Roh, 2006;) depends on Ring1 proteins or on another E3 ligase (s). The fact that linker histone H1 association is facilitated by H2AK119Ub1 (Jason, 2005) may also contribute to transcriptional repression, and one of the consequences of H2A deubiquitylation is the phosphorylation and release of histone H1 from nucleosomes (Zhu, 2007).…”

“…This bulky modification occurs at the proximity of the binding site of linker histones, facilitating the association of histone H1 to nucleosomes (Jason, 2005). During the isolation of a complex (es) responsible for H2A monoubiquitilation, a PRC1-like (PRC1l) complex, consisting of RING1A, RING1B, BMI1/PCGF4 and PH2/PHC2 was identified .…”