CAF-1 and Rtt101p function within the replication-coupled chromatin assembly network to promote H4 K16ac, preventing ectopic silencing

Abstract

Replication-coupled chromatin assembly is achieved by a network of alternate pathways containing different chromatin assembly factors and histone-modifying enzymes that coordinate deposition of nucleosomes at the replication fork. Here we describe the organization of a CAF-1-dependent pathway in Saccharomyces cerevisiae that regulates acetylation of histone H4 K16. We demonstrate factors that function in this CAF-1-dependent pathway are important for preventing establishment of silenced states at inappropriate genomic sites using a crippled HMR locus as a model, while factors specific to other assembly pathways do not. This CAF-1-dependent pathway required the cullin Rtt101p, but was functionally distinct from an alternate pathway involving Rtt101p-dependent ubiquitination of histone H3 and the chromatin assembly factor Rtt106p. A major implication from this work is that cells have the inherent ability to create different chromatin modification patterns during DNA replication via differential processing and deposition of histones by distinct chromatin assembly pathways within the network.

Detection and quantification of the histone code in the fungal genusAspergillus

“…For histone protein H3, we observed the conservation of all lysine residues in human, yeast, and Aspergilli, except for two additional lysine residues at positions 121 and 125 in S. cerevisiae (Figure 1). On these two residues, Cul8/Rtt101, an enzyme that is thought to be specific to S. cerevisiae as no homolog has yet been found in human or other fungi, can introduce histone ubiquitylation and thereby regulate nucleosome assembly (45,46). We similarly observed limited variation in the N-terminal tail of histone protein H4 and the global core domains of H2A and H2B, and most of the polymorphism occurs on amino acid residues that are not known to carry any histone PTM.…”

Chromatin assembly factor-1 preserves genome stability in ctf4∆ cells by promoting sister chromatid cohesion

“…Cohesin enrichment is enhanced genome-wide in response to DSB induction and this enrichment at undamaged sites globally tight-ens sister chromatid cohesion [ 124 ] [ 130 ] [ 131 ]. Taking into account that CAF-1 plays multiple roles in maintaining genome stability [ 3 ] [ 6 ] [ 7 ] [ 11 ] [ 13 ] [ 96 ] [ 98 ] [ 122 ] and that the absence of Cac1 in both wild-type and ctf4 Δ cells increased Rad52 foci ( Figure 3A , left), Rfa1 foci ( Figure 3A , right) [ 7 ], and γH2AX ( Figure 3B ), a possible explanation could be that DNA-damage-induced cohesion establishment was the source of the high cohesin level arising in absence of Cac1. Because Chk1, which mediates the DNA damage response in parallel with RAD53 , is a key component of the damage-induced cohesion establishment pathway required for the generation of damage-induced cohesion [ 134 ], we analyzed cohesin levels in absence of Chk1 in WT and cac1 Δ cells.…”

“…In absence of Ctf4, multiple cohesion establishment pathways are affected (red cross) leading to major defects in SCC establishment. In absence of CAF-1 fewer nucleosomes are deposited on replicated DNA, generating fewer but longer Okazaki fragments [ 140 ], leading to increased inter-nucleosome spacing in nascent chromatin and inappropriate epigenetic states [ 13 ] [ 148 ]. In these conditions, the nucleosome assembly function of CAF-1 is required to maintain SCC and efficient cell growth in yeast affected in cohesion.…”

“…In budding yeast, CAF-1 consists of three subunits called Cac1, Cac2, and Cac3, which differ in their ability to bind H3-H4. Deletion of any CAF-1 subunits is viable but leads to multiple defects including replisome dysfunction and DNA damage sensitivity [ 3 ] [ 7 ] [ 8 ] [ 9 ] [ 10 ] [ 11 ] [ 12 ] [ 13 ]. The ability of CAF-1 to deposit H3-H4 onto replicating DNA depends on its physical interaction with PCNA, a processivity factor for DNA polymerases, which is localized at the sites of DNA synthesis during replication and repair [ 14 ] [ 15 ] [ 16 ] [ 17 ] [ 18 ].…”

The nucleoporin Nup170 mediates subtelomeric gene silencing through the Ctf18-RFC complex and PCNA

“…Therefore, an intricate balance of PCNA levels on DNA that is tailored for its various activities is necessary for it to perform its functions. PCNA binds to chromatin assembly factor 1 (CAF-1) which recruits histones to accomplish DNA replication- or DNA repair-coupled nucleosome assembly (Gaillard et al, 1996; Young et al, 2020). PCNA accumulates and recruits CAF-1 to the damage site to facilitate nucleosome assembly during DNA damage repair (Essers et al, 2005; Gaillard et al, 1996; Moggs et al, 2000).…”