Replication-Coupled Nucleosome Assembly in the Passage of Epigenetic Information and Cell Identity

Abstract

During S phase, replicated DNA must be assembled into nucleosomes using both newly synthesized and parental histones in a process that is tightly coupled to DNA replication. This DNA replication-coupled process is regulated by multitude of histone chaperones as well as by histone-modifying enzymes. In recent years novel insights into nucleosome assembly of new H3-H4 tetramers have been gained through studies on the classical histone chaperone CAF-1 and the identification of novel factors involved in this process. Moreover, in vitro reconstitution of chromatin replication has shed light on nucleosome assembly of parental H3-H4, a process that remains elusive. Finally, recent studies have revealed that the replication-coupled nucleosome assembly is important for the determination and maintenance of cell fate in multicellular organisms.

Keywords: DNA replication; cell fate maintenance; epigenetic inheritance; histone modifications; nucleosome assembly.

Figure 1. Nucleosome assembly at DNA replication forks

Ahead of the replication fork, the MCM helicase unwinds the parental double-stranded DNA. The histone chaperone FACT, which interacts with MCM, presumably helps the advancement of the replication fork by disassembling parental nucleosomes. Parental H3–H4 tetramers could then be transferred to Mcm2 and FACT. However, how parental H3–H4 tetramers are shuttled to replicating DNA behind DNA replication forks and how they are deposited is currently unknown. The leading strand is synthesized in the same direction as the advancing replication fork by the DNA polymerase ε, which is in close contact with the CMG helicase. Each of the Okazaki fragments at the lagging strand requires an RNA primer first synthesized by DNA polymerase α. The remainder of the Okazaki fragment is synthesized by DNA polymerase δ, accompanied by PCNA. Newly synthesized H3–H4 dimers are escorted by Asf1 and transferred to downstream histone chaperones including CAF-1, which interacts with PCNA and DNA to deposit H3–H4 tetramers, the first step in nucleosome assembly of new H3–H4. Additionally, in yeast cells Asf1 transfers newly synthesized H3–H4 dimers to Rtt106, which deposits them cooperatively with FACT. RPA binds to ssDNA predominantly at the lagging strand template and regulates nucleosome assembly at the adjoining dsDNA by interacting with H3–H4 and several histone chaperones.

Figure 2. Chromatin maturation after DNA replication

Mature chromatin has a characteristic nucleosome pattern at gene regulatory elements such as the nucleosome-free region (NFR) upstream of the transcription start site (TSS) and the strongly positioned nucleosome (+1) immediately downstream of the TSS. The coding sequence of active genes also displays tight nucleosome spacing. In Saccharomyces cerevisiae, the NFR and the +1 nucleosome are rapidly restored soon after DNA replication by the action of transcription factors (TFs) and chromatin remodelers (left panel). The nucleosome phasing observed at gene bodies is restored later and is dependent on the transcription rate of a given gene and the activity of the HIR complex and chromatin remodelers. In contrast, the NFRs and the +1 nucleosomes are lost during the replication-coupled nucleosome assembly in Drosophila melanogaster (right panel). The nucleosome landscape at active regions is restored, more than an hour after DNA replication by TFs and chromatin remodelers. This may create a window of opportunity for alterations in chromatin state and cell identity.