Dynamic Assembly and Disassembly of the Human DNA Polymerase ? Holoenzyme on the Genome In Vivo.

ABSTRACT: Human DNA polymerase delta (Pol ?) forms a holoenzyme complex with the DNA sliding clamp proliferating cell nuclear antigen (PCNA) to perform its essential roles in genome replication. Here, we utilize live-cell single-molecule tracking to monitor Pol ? holoenzyme interaction with the genome in real time. We find holoenzyme assembly and disassembly in vivo are highly dynamic and ordered. PCNA generally loads onto the genome before Pol ?. Once assembled, the holoenzyme has a relatively short lifetime on the genome, implying multiple Pol ? binding events may be needed to synthesize an Okazaki fragment. During disassembly, Pol ? dissociation generally precedes PCNA unloading. We also find that Pol ? p125, the catalytic subunit of the holoenzyme, is maintained at a constant cellular level, indicating an active mechanism for control of Pol ? levels in vivo. Collectively, our studies reveal that Pol ? holoenzyme assembly and disassembly follow a predominant pathway in vivo; however, alternate pathways are observed.

SUBMITTER: Drosopoulos WC 

PROVIDER: S-EPMC7597369 | biostudies-literature | 2020 Feb

REPOSITORIES: biostudies-literature