Non-histone protein

About: Non-histone protein is a research topic. Over the lifetime, 1352 publications have been published within this topic receiving 75203 citations.

Chromatin structure: a repeating unit of histones and DNA.

Abstract: ciations of the histones in chromatin but says nothing of details, such as whether the F2A1 and F3 pair, which occurs as an (F2Al)2(F3)2 tetramer in solution, also occurs as a tetramer in chromatin. The most direct evidence for an (F2Al)2(F3)2 tetramer in chromatin is that a complex formed from tetramers, F2A2-F2B oligomers, and DNA gives the same x-ray pattern as chromatin (Fig. 4, upper two traces). Tetramers and F2A2-F2B oligomers are both required to give the x-ray pattern (Fig. 4, lower two traces), but Fl is not-in keeping with previous observations (3, 23 ) that removing Fl from chromatin does not affect the x-ray pattern. Further implications of these results are discussed in the accompanying article (24). We are currently studying associations of the histones in chromatin by cross-linking. There are two difficulties that do not arise in experiments on the histones in solution: the amino side chains are involved in salt linkages with the phosphate groups of DNA and are thus less available for chemical modification; and the presence of five rather than two histones complicates identification of products from molecular weights. -Preliminary results do show less cross-linking of histones in chromatin than in solution, but crosslinked products up to pentamers are readily observed and call for further investigation.

Mechanisms of Polycomb gene silencing: knowns and unknowns

Abstract: Polycomb proteins form chromatin-modifying complexes that implement transcriptional silencing in higher eukaryotes. Hundreds of genes are silenced by Polycomb proteins, including dozens of genes that encode crucial developmental regulators in organisms ranging from plants to humans. Two main families of complexes, called Polycomb repressive complex 1 (PRC1) and PRC2, are targeted to repressed regions. Recent studies have advanced our understanding of these complexes, including their potential mechanisms of gene silencing, the roles of chromatin modifications, their means of delivery to target genes and the functional distinctions among variant complexes. Emerging concepts include the existence of a Polycomb barrier to transcription elongation and the involvement of non-coding RNAs in the targeting of Polycomb complexes. These findings have an impact on the epigenetic programming of gene expression in many biological systems.

Phase separation drives heterochromatin domain formation

Abstract: HP1a can nucleate into foci that display liquid properties during the early stages of heterochromatin domain formation in Drosophila embryos, suggesting that the repressive action of heterochromatin may be mediated in part by emergent properties of phase separation. The gene-silencing action of heterochromatin is thought to arise from the spread of proteins such as HP1 that compact the underlying chromatin and recruit repressors. Two papers in this issue demonstrate that HP1α has the ability to form phase-separated droplets. Gary Karpen and colleagues show that HP1α can nucleate into foci that display liquid properties during the early stages of heterochromatin domain formation in Drosophila embryos. Geeta Narlikar and colleagues demonstrate that human HP1α protein also forms phase-separated droplets. Phosphorylation or DNA binding promotes the physical partitioning of HP1α out of the soluble aqueous phase into droplets. These related findings suggest that the repressive action of heterochromatin may be in part mediated by the phase separation of HP1, with the droplets being initiated or dissolved by various ligands depending on nuclear context. Constitutive heterochromatin is an important component of eukaryotic genomes that has essential roles in nuclear architecture, DNA repair and genome stability1, and silencing of transposon and gene expression2. Heterochromatin is highly enriched for repetitive sequences, and is defined epigenetically by methylation of histone H3 at lysine 9 and recruitment of its binding partner heterochromatin protein 1 (HP1). A prevalent view of heterochromatic silencing is that these and associated factors lead to chromatin compaction, resulting in steric exclusion of regulatory proteins such as RNA polymerase from the underlying DNA3. However, compaction alone does not account for the formation of distinct, multi-chromosomal, membrane-less heterochromatin domains within the nucleus, fast diffusion of proteins inside the domain, and other dynamic features of heterochromatin. Here we present data that support an alternative hypothesis: that the formation of heterochromatin domains is mediated by phase separation, a phenomenon that gives rise to diverse non-membrane-bound nuclear, cytoplasmic and extracellular compartments4. We show that Drosophila HP1a protein undergoes liquid–liquid demixing in vitro, and nucleates into foci that display liquid properties during the first stages of heterochromatin domain formation in early Drosophila embryos. Furthermore, in both Drosophila and mammalian cells, heterochromatin domains exhibit dynamics that are characteristic of liquid phase-separation, including sensitivity to the disruption of weak hydrophobic interactions, and reduced diffusion, increased coordinated movement and inert probe exclusion at the domain boundary. We conclude that heterochromatic domains form via phase separation, and mature into a structure that includes liquid and stable compartments. We propose that emergent biophysical properties associated with phase-separated systems are critical to understanding the unusual behaviours of heterochromatin, and how chromatin domains in general regulate essential nuclear functions.

Liquid droplet formation by HP1α suggests a role for phase separation in heterochromatin

Abstract: Gene silencing by heterochromatin is proposed to occur in part as a result of the ability of heterochromatin protein 1 (HP1) proteins to spread across large regions of the genome, compact the underlying chromatin and recruit diverse ligands. Here we identify a new property of the human HP1α protein: the ability to form phase-separated droplets. While unmodified HP1α is soluble, either phosphorylation of its N-terminal extension or DNA binding promotes the formation of phase-separated droplets. Phosphorylation-driven phase separation can be promoted or reversed by specific HP1α ligands. Known components of heterochromatin such as nucleosomes and DNA preferentially partition into the HP1α droplets, but molecules such as the transcription factor TFIIB show no preference. Using a single-molecule DNA curtain assay, we find that both unmodified and phosphorylated HP1α induce rapid compaction of DNA strands into puncta, although with different characteristics. We show by direct protein delivery into mammalian cells that an HP1α mutant incapable of phase separation in vitro forms smaller and fewer nuclear puncta than phosphorylated HP1α. These findings suggest that heterochromatin-mediated gene silencing may occur in part through sequestration of compacted chromatin in phase-separated HP1 droplets, which are dissolved or formed by specific ligands on the basis of nuclear context.