Mechanistic insights into histone deposition and nucleosome assembly by the chromatin assembly factor-1

Abstract

Eukaryotic chromatin is a highly dynamic structure with essential roles in virtually all DNA-dependent cellular processes. Nucleosomes are a barrier to DNA access, and during DNA replication, they are disassembled ahead of the replication machinery (the replisome) and reassembled following its passage. The Histone chaperone Chromatin Assembly Factor-1 (CAF-1) interacts with the replisome and deposits H3-H4 directly onto newly synthesized DNA. Therefore, CAF-1 is important for the establishment and propagation of chromatin structure. The molecular mechanism by which CAF-1 mediates H3-H4 deposition has remained unclear. However, recent studies have revealed new insights into the architecture and stoichiometry of the trimeric CAF-1 complex and how it interacts with and deposits H3-H4 onto substrate DNA. The CAF-1 trimer binds to a single H3-H4 dimer, which induces a conformational rearrangement in CAF-1 promoting its interaction with substrate DNA. Two CAF-1•H3-H4 complexes co-associate on nucleosome-free DNA depositing (H3-H4)2 tetramers in the first step of nucleosome assembly. Here, we review the progress made in our understanding of CAF-1 structure, mechanism of action, and how CAF-1 contributes to chromatin dynamics during DNA replication.

Figure 1.

Histone chaperones orchestrate nucleosome assembly during DNA replication. Nucleosomes are removed ahead of the advancing MCM helicase complex and reassembled onto the daughter strands after the passage of the replisome. The transfer of (H3–H4)₂ tetramers occurs conservatively, i.e. is distributed as a single unit onto one of the two daughter strands. FACT, MCM2 and ASF1 are histone chaperones that might be involved here (23,24,30). De novo assembly of nucleosomes occurs primarily via CAF-1 which receives histone dimers from ASF1 and deposits (H3–H4)₂ tetramers through a mechanism described in the text. Other chaperones deposit H2A–H2B to complete recycled and de novo nucleosome assembly. The timely assembly of nucleosomes is critical for replication fork progression and lagging strand synthesis. After assembly, nucleosome positions are fine-tuned by ATP dependent chromatin remodelers (not shown) (7).

Figure 2.

Architecture of the CAF-1 complex: (A) domain arrangement of the large CAF-1 subunit in humans and yeast. The primary structures of both homologues are shown with selected protein-protein and protein-DNA interaction domains highlighted. Sequence disorder predictions (obtained from DISOPRED3 (56)) are shown. PIP—PCNA interacting peptide (34–36); SUMO - small ubiquitin like modifier (60); PxVxL is a HP1 (Heterochromatin protein) interacting sequence motif (61); The PEST sequence is associated with reduced intracellular half-life (32); KER sequence binds to DNA (55); ED sequence binds to histones and WHD (58); WHD—Winged helix domain (64). (B) Superimposition of Drosophila RbAp48/p55 (orange) bound to H3 (green, residues 1–11; PDB 2YBA) or H4 (blue, residues 30–43; PDB 2XYI). The WD40 blades of p55 are numbered, starting from the N-terminus. Note that alignment of the H4 peptide to nucleosomal H4 results in clashes of the nucleosomal H3–H4 pair with p55 (78–80). (C) Overall organization of the CAF-1 complex and interacting regions, including ASF1-H3–H4. Individual domains or regions are connected with arrows and represent only approximations, based on XL-MS, HX-MS and mutagenesis experiments. Individual WD40 blades of Cac2 and Cac3 are indicated and were predicted by Phyre2. ASF1delivers H3–H4 to CAF-1 but is not part of the complex that carries out deposition. See text for details (26,55,58,62,63,83). (D) Ab initio model calculated from SAXS data. The predicted subunit regions within CAF-1 are labeled. The dimensions for the complex, and subunits are noted (62).

Figure 3.

Model for CAF-1 mediated nucleosome assembly. In absence of histones, the C-terminal WHD is inaccessible due to sequestration by the ED domain. ASF1 transfers a single H3–H4 dimer to CAF-1, resulting in the liberation of the WHD. Two CAF-1•H3–H4 complexes associate in close proximity to each other, mediated by PCNA–CAF-1 and DNA–CAF-1 contacts. The transient association of two CAF-1•H3–H4 complexes on DNA allows for H3-H3 contacts to form and the two histone chaperone complexes concertedly deposit one (H3–H4)₂ tetramer onto the DNA prior to being released from the DNA. During the second assembly step, H2A–H2B histone chaperones mediate H2A–H2B deposition onto the preexisting tetramer forming full nucleosomes. For details see text.