Histone modification governs the cell cycle regulation of a replication-independent chromatin assembly pathway in Saccharomyces cerevisiae

Abstract

We describe a replication-independent, cell cycle-regulated chromatin assembly pathway in budding yeast. The activity of this pathway is low in S phase extracts but is very high in G 2 , M, and G 1 cell extracts, with peak activity in late M͞early G 1 . The cell cycle regulation of this pathway requires a specific pattern of posttranslational modification of histones H3 and͞or H4, which is distinct for H3͞H4 present in S phase versus M and G 1 phase cell extracts. Histone H3͞H4 modification is therefore important for the reciprocal control of replication-dependent and -independent chromatin assembly pathways during the cell cycle.Under physiological conditions nucleosome assembly is mediated by proteins called chromatin assembly factors (CAFs) (1-5). CAFs operate in two general pathways of nucleosome reconstitution in vivo, one coupled to DNA replication, and one that occurs independently of replication. The ''replicationdependent'' pathway is cell cycle regulated, being maximally active in S phase. Although this pathway accounts for the bulk of chromatin assembly in dividing cells, it seems likely that during normal cell division nucleosome deposition also occurs by a ''replication-independent'' mechanism. Chromatin remodeling associated with transcriptional regulation can involve nucleosome deposition that is independent of DNA synthesis (6) and therefore perhaps involves replicationindependent assembly factors. When nucleosomes are lost during G 2 , M, and G 1 as a result of histone degradation, new nucleosome deposition is likely to occur before the next S phase (7), and this reaction also potentially involves replication-independent CAFs. Although these observations suggest significant physiological functions for replication-independent CAFs in dividing cells, the biochemical properties of such factors, and the regulation of replication-independent assembly pathways in relation to global changes in cellular metabolism, are poorly understood. We are exploring the regulation of replicationindependent chromatin assembly by a biochemical approach in budding yeast (8, 9). Here we focus on the cell cycle regulation of replication-independent chromatin assembly in mitotically dividing cells. We have discovered a replication-independent assembly pathway that is cell cycle regulated in yeast. The activity pattern of this pathway mirrors that of the replicationdependent pathway responsible for bulk assembly in dividing cells, that is, replication-independent assembly is repressed in S phase, during which time bulk replication-coupled assembly is activated (2, 10). We show that posttranslational modification of histones H3 and͞or H4 plays an important role in the cell cycle control of assembly by this pathway. Posttranslational modification of histones H3 and H4 is also associated with the activation of replication-coupled chromatin assembly during S phase (2, 10). The regulation of histone H3 and͞or H4 is therefore an important component of a reciprocal control mechanism that represses replication-independent...

Global Control of Histone Modification by the Anaphase-Promoting Complex

“…Against a background of degradation of free hyperacetylated histones, this mechanism would drive global replacement of hyper-with hypoacetylated histones. This idea is consistent with our evidence that chromatin assembly in vitro is responsive to the state of histone modification (3,50) and that extracts from APC10 (this study) and APC5 (25) mutants have a defect in assembly activity. In order to critically test this possibility, it will be necessary to identify the assembly factors that are active in the crude yeast assembly system and to test their use of modified versus unmodified histones during active proliferation and in G 0 .…”

“…We previously discovered that mutation of the conserved APC5 subunit of the APC inhibits replication-independent chromatin assembly in a yeast extract (25). The reaction in this system is also sensitive to mutation of amino-terminal lysines in H4 that can be acetylated in vivo, and H3 and H4 from G 2 /M cells are better substrates for assembly in the extract than are H3 and H4 from S-phase cells (3,46,50). It follows that a defect in histone metabolism may underlie the assembly defect of extract from apc5 mutant cells.…”

Histone Variants and Epigenetics

“…Work in budding yeast revealed that none of the subunits of complexes involved in RC assembly in vitro is essential for growth, suggesting that in vivo, there are redundant mechanisms for RC assembly. The fact that much of yeast chromatin is assembled in a replication-independent (RI) manner (Altheim and Schultz 1999) provides a rationale for this evident redundancy. As we shall see, histone variants are typically deposited by RI nucleosome assembly.…”

Replication-Independent Assembly of Nucleosome Arrays in a Novel Yeast Chromatin Reconstitution System Involves Antisilencing Factor Asf1p and Chromodomain Protein Chd1p

“…Alternatively, it has been demonstrated that nucleosomes can be assembled or reassembled independently of DNA replication. Replication-independent (RI) assembly is not limited to the S phase but occurs continually throughout the cell cycle (2,4), perhaps functioning as a backup to RC assembly (44). RI assembly can introduce specific histone variants, such as the H3 variant Cid, at centrosomes (1) or histone H3.3 in transcriptionally active regions of the genome (2).…”