Structure and function of the histone chaperone FACT - Resolving FACTual issues

Abstract

FAcilitates Chromatin Transcription (FACT) has been considered essential for transcription through chromatin mostly based on cell-free experiments. However, FACT inactivation in cells does not cause a significant reduction in transcription. Moreover, not all mammalian cells require FACT for viability. Here we synthesize information from different organisms to reveal the core function(s) of FACT and propose a model that reconciles the cell-free and cell-based observations. We describe FACT structure and nucleosomal interactions, and their roles in FACT-dependent transcription, replication and repair. The variable requirements for FACT among different tumor and non-tumor cells suggest that various FACT-dependent processes have significantly different levels of relative importance in different eukaryotic cells. We propose that the stability of chromatin, which might vary among different cell types, dictates these diverse requirements for FACT to support cell viability. Since tumor cells are among the most sensitive to FACT inhibition, this vulnerability could be exploited for cancer treatment.

Keywords: Cancer; Chromatin; Histone; SPT16; SSRP1; Transcription.

Figure 1.. Schematic structures of human (h), yeast (y), and plant (p) FACT complexes.

NTD – N-terminal domain, DD – dimerization domain, MD – middle domain, CTD – C-terminal intrinsically disordered domain, IDD – intrinsically disordered domain, HMG - HMG box domain. Structural data are available for filled domains. Additional features and some posttranslational modifications are shown for human FACT subunits: blue crescent - nuclear localization signal, yellow circles - phosphorylation sites, pale purple circles - acetylated lysines.

Figure 2.. Schematic representation of interactions of FACT subunits domains with components of nucleosome.

An assembled nucleosome is shown in the upper left corner. A color code is used to indicate histones and histone tails. Black solid arrows – established physical interactions, black dashed arrows – hydrophobic and electrostatic interactions. Grey arrows - less well established interactions.

Figure 3.. Mechanism of FACT action during transcription through a nucleosome.

As Pol II approaches a nucleosome (complex 1), nucleosomal DNA is partially uncoiled from the octamer, exposing DNA-binding surface of the promoter-proximal H2A/H2B dimer (2); FACT interacts with DNA-binding surface of the exposed dimer, preventing re-coiling of nucleosomal on the octamer and formation of a non-productive complex 2’. Then a transient, small intranucleosomal DNA Ø-loop is formed at the position +49 (3); Pol II can transcribe further only after displacement of the promoter-distal end of nucleosomal DNA by Pol II, exposing DNA- binding surface of the promoter-distal dimer (4). Here ~100-bp DNA region is uncoiled from the octamer, likely resulting in a loss of an H2A/H2B dimer. As Pol II continues transcription, the DNA-histone contacts upstream of the enzyme serve as an anchor to recover the nucleosome behind Pol II (5–6). The dimer is recovered only after completion of transcription (6). FACT transiently interacts with the nucleosome-bound H2A/H2B dimers during transcription (intermediates 2 and 4), transiently prevents DNA recoiling on the histone octamer and affects the rate of transcription through chromatin. FACT also facilitates nucleosome survival and could facilitate loss of a dimer during the 4 -> 5 transition.