The conserved histone chaperone Spt6 is strongly required for DNA replication and genome stability

A fluorescent assay for cryptic transcription inSaccharomyces cerevisiaereveals novel insights into factors that stabilize chromatin structure on newly replicated chromatin

“…Figures 5A and S9 show that a G1 arrest before the shift to the non-permissive temperature partially suppressed the cryptic transcription phenotype, suggesting that some of the chromatin disruption in the absence of FACT was due to DNA replication. Similar results were observed in a temperature-sensitive mutant of SPT6 (Figures 5B and S10), which encodes a histone chaperone recently implicated in DNA replication (Miller and Winston 2023). Because both SPT6 and SPT16 mutants were grown on dextrose, which represses transcription from the upstream GAL1 promoter, the incomplete rescue implies that depletion of FACT or Spt6 results in chromatin disruption in the absence of DNA replication and transcription.…”

“…Multiple factors play dual functions in stabilizing chromatin structure on both transcribed and newly replicated DNA, including Asf1, Rtt106, Spt6, and FACT (Wittmeyer et al 1999;Schlesinger and Formosa 2000;O'Donnell et al 2004;Zhou and S-F Wang 2004;Groth et al 2005Groth et al , 2007Franco et al 2005;Schulz and Tyler 2006;VanDemark et al 2006;Sanematsu et al 2006;Han et al 2007;Li et al 2008a;Grigsby et al 2009;Abe et al 2011;Ishikawa et al 2011;Zunder et al 2012;Klimovskaia et al 2014;Yang et al 2016;Safaric et al 2022;Nakagawa et al 2022;Miller and Winston 2023), and we used our reporter assay to test whether some of the chromatin disruption that occurs in the absence of these factors is due to DNA replication.…”

“…Although transcription from an upstream promoter is required for the activation of cryptic promoters in some mutants, it is not strictly needed, including in ASF1, SPT6, SPT16, and SPT21 mutants (Cheung et al 2008), suggesting that a process other than transcription can destabilize nucleosomes. An obvious candidate is DNA replication, which is inherently disruptive to chromatin structure, and indeed, many replication-coupled factors, including Asf1, Rtt106, Spt6, and Spt16, are suppressors of cryptic transcription (Wittmeyer et al 1999;Schlesinger and Formosa 2000;O'Donnell et al 2004;Zhou and S-F Wang 2004;Groth et al 2005Groth et al , 2007Franco et al 2005;Schulz and Tyler 2006;VanDemark et al 2006;Sanematsu et al 2006;Han et al 2007;Li et al 2008a;Grigsby et al 2009;Abe et al 2011;Ishikawa et al 2011;Zunder et al 2012;Klimovskaia et al 2014;Yang et al 2016;Safaric et al 2022;Nakagawa et al 2022;Miller and Winston 2023). However, these proteins also have transcription-linked functions (Orphanides et al 1998;Mason and Struhl 2003;Adkins et al 2004;Biswas et al 2005;Imbeault et al 2008;Birch et al 2009;Hsieh et al 2013), and whether DNA replication is required for cryptic transcription following loss of these suppressor proteins has never been tested.…”

The MCM2-7 Complex: Roles beyond DNA Unwinding

“…In budding yeast, FACT interacts with the MCM2 subunit, while, in higher eukaryotes, FACT may have a different interacting partner within the complex [126,127,[129][130][131]. Indeed, the FACT complex, originally characterized for its role in transcription, seems to play critical roles in assuring the efficiency of DNA replication through different kinds of chromatin, suggesting that the coupling of DNA replication with histone repositioning improves the efficiency of DNA replication [128,129,[131][132][133]. Collectively, the data indicate that the MCM2-7 complex at the core of the active replication helicase coordinates histone deposition adjacent to the replication fork.…”