Hst3 is turned over by a replication stress-responsive SCF ^Cdc4 phospho-degron

Abstract

Hst3 is the histone deacetylase that removes histone H3K56 acetylation. H3K56 acetylation is a cell-cycle-and damage-regulated chromatin marker, and proper regulation of H3K56 acetylation is important for replication, genomic stability, chromatin assembly, and the response to and recovery from DNA damage. Understanding the regulation of enzymes that regulate H3K56 acetylation is of great interest, because the loss of H3K56 acetylation leads to genomic instability. HST3 is controlled at both the transcriptional and posttranscriptional level. Here, we show that Hst3 is targeted for turnover by the ubiquitin ligase SCF Cdc4 after phosphorylation of a multisite degron. In addition, we find that Hst3 turnover increases in response to replication stress in a Rad53-dependent way. Turnover of Hst3 is promoted by Mck1 activity in both conditions. The Hst3 degron contains two canonical Cdc4 phospho-degrons, and the phosphorylation of each of these is required for efficient turnover both in an unperturbed cell cycle and in response to replication stress.H istone H3K56 acetylation is a cell-cycle-and damageregulated histone modification (1). In Saccharomyces cerevisiae, K56 acetylation is important for replication, genomic stability, chromatin assembly, and the response and recovery from DNA damage (1-6). Because of the critical nature of this chromatin mark, the enzymes that control it are themselves very important.In the budding yeast S. cerevisiae, Hst3 is a sirtuin histone deacetylase that, along with its close homolog Hst4, removes K56 acetylation (5, 7, 8). Overexpression of Hst3 or deletion of Hst3, along with related deacetylase Hst4, sensitizes cells to replication stress, suggesting that cells must control Hst3 levels precisely (5, 7). Hst3 is regulated both transcriptionally and by protein turnover. During an unperturbed cell cycle, HST3 transcript levels cycle, and Hst3 protein turns over very quickly (5,9). In response to DNA damage, transcription of HST3 is turned down, and Hst3 protein turnover is even faster (5,8,(10)(11)(12).Many cell-cycle-regulated proteins are targeted for proteasomal degradation after ubiquitination by a member of the SCF E3 ubiquitin ligase family. SCF ligases are multisubunit ubiquitin ligases composed of Skp1, a cullin (Cdc53), a RING-finger protein (Rbx1), and an F-box protein (13). The F-box protein is the substrate recognition module that confers substrate specificity to SCF ligases, with different F-box proteins recognizing different sets of substrates. The essential F-box protein Cdc4 regulates many cell-cycle-regulated proteins after their phosphorylation, including Sic1, Eco1, Cln3, and many others (14-19).Here, we have characterized the mechanism of Hst3 protein turnover. We find that Hst3 is targeted for degradation by SCF Cdc4 through a multisite phospho-degron. We find that Hst3 turnover is increased in response to replication stress in a Rad53-dependent way. Finally, we show that stabilizing Hst3 leads to misregulation of K56 acetylation in response to DNA damage and that...

Epigenetic Control of Expression Homeostasis during Replication Is Stabilized by the Replication Checkpoint

“…It was previously reported that Mec1 targets the H3K56ac deacetylase Hst3 to degradation (Edenberg et al, 2014). We reasoned that this degradation could explain the reduced H3K56ac and loss of homeostasis.…”

“…We reasoned that this degradation could explain the reduced H3K56ac and loss of homeostasis. To this end, we considered Hst3D97, an Hst3 allele that is resistant to the Mec1-dependent phosphorylation that leads to its degradation (Edenberg et al, 2014). This allele, however, had no effect on expression homeostasis ( Figures S5K-S5M).…”

Prb1 Protease Activity Is Required for Its Recognition by the F-Box Protein Saf1

“…6,7 Degrons are usually N- or C-terminal extensions outside the catalytic domain and function when fused to ectopic substrates. 8,9 …”

Nicotinamide Suppresses the DNA Damage Sensitivity of Saccharomyces cerevisiae Independently of Sirtuin Deacetylases

“…Four of the five sirtuins have reported roles in responding to or preventing enhanced accumulation of DNA damage. Hst3 and Hst4 deacetylate histone H3 on K56, which is deposited during DNA replication, and this deacetylation is prevented in the presence of DNA damage (Celic et al 2006;Maas et al 2006;Miller et al 2006;Edenberg et al 2014b). Levels of Hst3 and Hst4 are tightly controlled (Thaminy et al 2007;Delgoshaie et al 2014;Edenberg et al 2014b) and their absence results in persistent H3 K56 acetylation.…”

“…Hst3 and Hst4 deacetylate histone H3 on K56, which is deposited during DNA replication, and this deacetylation is prevented in the presence of DNA damage (Celic et al 2006;Maas et al 2006;Miller et al 2006;Edenberg et al 2014b). Levels of Hst3 and Hst4 are tightly controlled (Thaminy et al 2007;Delgoshaie et al 2014;Edenberg et al 2014b) and their absence results in persistent H3 K56 acetylation. Deregulation of H3 K56 acetylation is associated with spontaneous activation of the DNA damage response and defects in DNA repair (Celic et al 2006(Celic et al , 2008Maas et al 2006;Muñoz-Galván et al 2013;Che et al 2015;Simoneau et al 2015).…”