FACT Facilitates Transcription-Dependent Nucleosome Alteration

Abstract

The FACT (facilitates chromatin transcription) complex is required for transcript elongation through nucleosomes by RNA polymerase II (Pol II) in vitro. Here, we show that FACT facilitates Pol II-driven transcription by destabilizing nucleosomal structure so that one histone H2A-H2B dimer is removed during enzyme passage. We also demonstrate that FACT possesses intrinsic histone chaperone activity and can deposit core histones onto DNA. Importantly, FACT activity requires both of its constituent subunits and is dependent on the highly acidic C terminus of its larger subunit, Spt16. These findings define the mechanism by which Pol II can transcribe through chromatin without disrupting its epigenetic status.

Chromatin disassembly and reassembly during DNA repair

“…For example, chromatin is disassembled from some yeast gene promoters during transcriptional activation by the H3/H4 chaperone Asf1 [32]. Another histone chaperone, FACT (facilitates chromatin transcription), mediates the transient disassembly and reassembly of histones H2A/ H2B within open reading frames of genes to facilitate passage of RNA polymerase II [33]. Similarly, the histone H3/H4 chaperone Spt6 mediates chromatin reassembly within open reading frames following RNA polymerase II passage in yeast [34] and the reassembly of promoter nucleosomes during transcriptional repression [35].…”

Coupling caspase cleavage and ubiquitin–proteasome-dependent degradation of SSRP1 during apoptosis

“…Immunoblot showed no detectable decrease of SSRP1 even after 24 h (Figure 1a, lanes 4,8,12), suggesting that this is a very stable protein. In contrast, p21 levels decreased dramatically 3 h post-treatment (lanes 2,6,10) confirming the efficiency of the CHX treatment. SSRP1 stability was confirmed using other translation inhibitors (puromycin, emetin) and by pulse-chase experiments (data not shown).…”

“…17,22 However, human SSRP1 is unable to bind mononucleosomes by itself and requires interaction with Spt16, 6 which occurs via the SSRP1 N-terminal. 13 These in vitro data suggest that the N-terminal part of SSRP1 is critical for its interaction with nucleosomes.…”

“…5 Human FACT promotes nucleosome disassembly in vitro by selectively removing one histone H2A-H2B dimer. 6,7 Interestingly, FACT appears to reassemble nucleosomes after the passage of the RNA polymerase II, reconstituting the repressive state of chromatin. 6 Remarkably, ATP is not required, suggesting that the mechanism by which FACT remodels nucleosomes differs fundamentally from other chromatin remodeling factors such as SWI/SNF.…”

“…6,7 Interestingly, FACT appears to reassemble nucleosomes after the passage of the RNA polymerase II, reconstituting the repressive state of chromatin. 6 Remarkably, ATP is not required, suggesting that the mechanism by which FACT remodels nucleosomes differs fundamentally from other chromatin remodeling factors such as SWI/SNF. 8 In yeast and xenopus, FACT is directly involved in DNA replication.…”