Chromatin Constrains the Initiation and Elongation of DNA Replication

Sujan Devbhandari; Jieqing Jiang; Charanya Kumar; Iestyn Whitehouse; Dirk Remus

doi:10.1016/j.molcel.2016.10.035

Chromatin Constrains the Initiation and Elongation of DNA Replication

Mol Cell. 2017 Jan 5;65(1):131-141. doi: 10.1016/j.molcel.2016.10.035. Epub 2016 Dec 15.

Authors

Sujan Devbhandari¹, Jieqing Jiang¹, Charanya Kumar², Iestyn Whitehouse², Dirk Remus³

Affiliations

¹ Molecular Biology Program, Memorial Sloan Kettering Cancer Center, 1275 York Avenue, New York, NY 10065, USA; Weill-Cornell Graduate School of Medical Sciences, New York, NY 10065, USA.
² Molecular Biology Program, Memorial Sloan Kettering Cancer Center, 1275 York Avenue, New York, NY 10065, USA.
³ Molecular Biology Program, Memorial Sloan Kettering Cancer Center, 1275 York Avenue, New York, NY 10065, USA. Electronic address: remusd@mskcc.org.

Abstract

Eukaryotic chromosomal DNA is faithfully replicated in a complex series of cell-cycle-regulated events that are incompletely understood. Here we report the reconstitution of DNA replication free in solution with purified proteins from the budding yeast Saccharomyces cerevisiae. The system recapitulates regulated bidirectional origin activation; synthesis of leading and lagging strands by the three replicative DNA polymerases Pol α, Pol δ, and Pol ε; and canonical maturation of Okazaki fragments into continuous daughter strands. We uncover a dual regulatory role for chromatin during DNA replication: promoting origin dependence and determining Okazaki fragment length by restricting Pol δ progression. This system thus provides a functional platform for the detailed mechanistic analysis of eukaryotic chromosome replication.

Keywords: DNA polymerase; DNA replication; McM2-7; ORC; Okazaki fragment; chromatin; lagging strand; leading strand; replication origin.

MeSH terms

Chromatin / genetics*
Chromatin / metabolism
DNA / genetics
DNA / metabolism
DNA Polymerase I / genetics
DNA Polymerase I / metabolism
DNA Polymerase II / genetics
DNA Polymerase II / metabolism
DNA Polymerase III / genetics
DNA Polymerase III / metabolism
DNA Replication*
DNA Topoisomerases, Type I / genetics
DNA Topoisomerases, Type I / metabolism
DNA Topoisomerases, Type II / genetics
DNA Topoisomerases, Type II / metabolism
DNA, Fungal / biosynthesis
DNA, Fungal / genetics*
Genotype
Humans
Nucleosomes / genetics*
Nucleosomes / metabolism
Phenotype
Proliferating Cell Nuclear Antigen / genetics
Proliferating Cell Nuclear Antigen / metabolism
Replication Origin*
Saccharomyces cerevisiae / genetics*
Saccharomyces cerevisiae / metabolism
Saccharomyces cerevisiae Proteins / genetics
Saccharomyces cerevisiae Proteins / metabolism
Time Factors

Substances

Chromatin
DNA, Fungal
Nucleosomes
Okazaki fragments
POL30 protein, S cerevisiae
Proliferating Cell Nuclear Antigen
Saccharomyces cerevisiae Proteins
DNA
DNA Polymerase I
DNA Polymerase II
DNA Polymerase III
DNA Topoisomerases, Type I
DNA Topoisomerases, Type II

Abstract

MeSH terms

Substances

Grants and funding