THE INTERDEPENDENCE OF DNA REPLICATION, DNA SEQUENCE AND CHROMATIN STRUCTURE IN SIMIAN VIRUS 40 CHROMOSOMES

Abstract

ABSTRACT. The interdependence of DNA replication, DNA sequence, and chromatin structure is readily seen in SV40 chromosomes where a unique DNA sequence is organized into nucleosomes whose structure and histone composition are indistinguishable from those of the host. During replication, SV40 DNA synthesis is carried out by DNA polymerase α in a semidiscontinuous manner via repeated initiation of Okazaki fragments that originate predominantly, if not exclusively, from retrograde templates. The metabolism of Okazaki fragments involves at least five transient intermediates, synthesis and excision of RNA primers, DNA polymerase α, and at least one protein co-factor. Following initiation of Okazaki fragments, nucleosomes rapidly appear on both arms of replication forks at about 125 bp from the 3′- and 5′ ends of long nascent DNA strands. Initiation of Okazaki fragments is promoted at many preferred DNA sequences within an “initiation zone” on the retrograde template. The size of this zone correlates with the average spacing of nucleosomes which are phased in a near-random fashion with respect to DNA sequence throughout the genome. Preferred DNA sequences are also involved in arresting the progress of replication forks in the termination region of the genome. These sites can account for the observed accumulation of DNA at 90% replication and may promote separation of sibling molecules. Purified DNA polymerase α is also arrested at preferred DNA sequences which may contribute to its ineffective-ness at completing synthesis of Okazaki fragments on purified DNA. These properties demonstrate that α-polymerase is sensitive to the structure of its template. Taken together, these observations support a functional role for DNA organ-ization and chromatin structure in the events at eukaryotic replication forks.