The role of FACT in making and breaking nucleosomes

Abstract

FACT is a roughly 180kDa heterodimeric protein complex important for managing the properties of chromatin in eukaryotic cells. Chromatin is a repressive barrier that plays an important role in protecting genomic DNA and regulating access to it. This barrier must be temporarily removed during transcription, replication, and repair, but it also must be rapidly restored to the original state afterwards. Further, the properties of chromatin are dynamic and must be adjusted as conditions dictate. FACT was identified as a factor that destabilizes nucleosomes in vitro, but it has now also been implicated as a central factor in the deposition of histones to form nucleosomes, as an exchange factor that swaps the histones within existing nucleosomes for variant forms, and as a tether that prevents histones from being displaced by the passage of RNA polymerases during transcription. FACT therefore plays central roles in building, maintaining, adjusting, and overcoming the chromatin barrier. This review summarizes recent results that have begun to reveal how FACT can promote what appear to be contradictory goals, using a simple set of binding activities to both enhance and diminish the stability of nucleosomes. This article is part of a Special Issue entitled: Histone chaperones and Chromatin assembly.

Figure 1

Domain structure of FACT proteins. The NT (N-terminal), D (dimerization), M (middle), C (C-terminal), and HMGB domains of S. cerevisiae Spt16, Pob3, and Nhp6 and human SSRP1 are diagrammed to scale. The acidic domains are colored red. SSRP1 has a serine-rich C-terminal extension not found in yeast or fungal FACT. The Spt16-NT domain has two lobes as indicated in cyan and blue, with the loop enclosing the conserved putative peptide-binding domain indicated in magenta [30, 31]. Ribbon diagrams of the domains whose crystallographic structures are known are shown, as is a surface representation of Nhp6 bound to DNA determined by NMR [90]. Structures were rendered in PyMOL using PDB files 3BIQ (Spt16-NT) [31], 3F5R (Pob3-NT/D) [35], 2GCL (Pob3-M) [12], and 1J5N (Nhp6-DNA) [90].

Figure 2

Model for nucleosome reorganizing activities Nhp6 binds to DNA at the surface of a canonical nucleosome, bending the DNA and destabilizing histone-DNA contacts [39]. Repeated binding-bending-release cycles within a short window of time render the nucleosome less stable and more suitable for reorganization by Spt16-Pob3. Reorganization involves disruption of the contacts between H2A–H2B and H3–H4, but the components of the nucleosome remain tethered together by FACT in a form in which the DNA is largely accessible. This dynamic form loses H2A–H2B dimers at a higher frequency than canonical nucleosomes do, but in the “global accessibility” model, dimer loss is an optional outcome of reorganization, not an obligate product. In the “dimer displacement” model FACT activity proceeds directly to displacement of an H2A–H2B dimer to form a hexasome. FACT might promote insertion of variant forms of H2A–H2B by chaperoning them and swapping them into either the reorganized form or hexasomes. The extended C-terminal tail of H2A that forms the docking domain that is partially responsible for dimer-tetramer stability in the core octamer is labeled; this domain may retain contact with H3–H4 during reorganization by bending at the point where it reaches the globular domain, unlike the rigid-body movement modeled here.

Figure 3

FACT’s potential physiological roles Canonical, repressive nucleosomes are indicated in red, with reorganized nucleosomes shown in green. In DNA replication (upper panel), FACT’s ability to promote nucleosome reorganization appears to contribute to activation of replication origins by providing access to the DNA, by promoting progression of the replication complex through chromatin by interacting with the MCM helicase and by destabilizing nucleosomes, by tethering of histones during replication, and by assisting the assembly/reassembly of nucleosomes after replication. Similar functions are proposed in transcription (lower panel). FACT collaborates at all steps with other histone chaperones, such as CAF-1, Asf1, Rtt106, and possibly Nap1, as well as ATP-dependent remodelers and chromatin modifying factors.