Dynamic loading and redistribution of the Mcm2-7 helicase complex through the cell cycle

EMBO J. 2015 Feb 12;34(4):531-43. doi: 10.15252/embj.201488307. Epub 2015 Jan 2.

Abstract

Eukaryotic replication origins are defined by the ORC-dependent loading of the Mcm2-7 helicase complex onto chromatin in G1. Paradoxically, there is a vast excess of Mcm2-7 relative to ORC assembled onto chromatin in G1. These excess Mcm2-7 complexes exhibit little co-localization with ORC or replication foci and can function as dormant origins. We dissected the mechanisms regulating the assembly and distribution of the Mcm2-7 complex in the Drosophila genome. We found that in the absence of cyclin E/Cdk2 activity, there was a 10-fold decrease in chromatin-associated Mcm2-7 relative to the levels found at the G1/S transition. The minimal amounts of Mcm2-7 loaded in the absence of cyclin E/Cdk2 activity were strictly localized to ORC binding sites. In contrast, cyclin E/Cdk2 activity was required for maximal loading of Mcm2-7 and a dramatic genome-wide reorganization of the distribution of Mcm2-7 that is shaped by active transcription. Thus, increasing cyclin E/Cdk2 activity over the course of G1 is not only critical for Mcm2-7 loading, but also for the distribution of the Mcm2-7 helicase prior to S-phase entry.

Keywords: DNA replication; Mcm2‐7; cell cycle; chromatin.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Blotting, Western
  • Cell Cycle / genetics
  • Cell Cycle / physiology*
  • Cells, Cultured
  • Drosophila
  • Drosophila Proteins / genetics
  • Drosophila Proteins / metabolism*
  • Fluorescent Antibody Technique
  • Minichromosome Maintenance Proteins / genetics
  • Minichromosome Maintenance Proteins / metabolism*
  • RNA Interference

Substances

  • Drosophila Proteins
  • Minichromosome Maintenance Proteins