Role of the second-largest subunit of DNA polymerase alpha in the interaction between the catalytic subunit and hyperphosphorylated retinoblastoma protein in late S phase

Biochim Biophys Acta. 2006 Sep;1764(9):1447-53. doi: 10.1016/j.bbapap.2006.06.015. Epub 2006 Jul 25.


DNA polymerase alpha (pol-alpha) is a heterotetrameric enzyme (p180-p68-p58-p48 in mouse) that is essential for the initiation of chain elongation during DNA replication. The catalytic (p180) and p68 subunits of pol-alpha are phosphorylated by Cdk-cyclin complexes, with p68 being hyperphosphorylated by cyclin-dependent kinases in G(2) phase of the cell cycle. The activity of Cdk2-cyclin A increases during late S phase and peaks in G(2) phase. We have now examined the role of p68 in the interaction between the catalytic subunit of pol-alpha and hyperphosphorylated retinoblastoma protein (ppRb) and in the stimulation of the polymerase activity of pol-alpha by ppRb. With the use of recombinant proteins, we found that nonphosphorylated p68 inhibited the stimulation of pol-alpha activity by ppRb, suggesting that p68 might impede the association of ppRb with p180. Phosphorylation of p68 by Cdk2-cyclin A greatly reduced its inhibitory effect. Immunofluorescence analysis also revealed that ppRb localized at sites of DNA replication specifically in late S phase. These results suggest that Cdk-cyclin A can phosphorylate pol-alpha which may result in a conformational change in pol-alpha facilitating its interaction with and activation by ppRb.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cyclin A / metabolism
  • Cyclin-Dependent Kinase 2 / metabolism
  • DNA Polymerase I / physiology*
  • HeLa Cells
  • Heterochromatin / metabolism
  • Humans
  • Mice
  • Phosphorylation
  • Protein Subunits / physiology*
  • Retinoblastoma Protein / metabolism*
  • S Phase / physiology*


  • Cyclin A
  • Heterochromatin
  • Protein Subunits
  • Retinoblastoma Protein
  • Cyclin-Dependent Kinase 2
  • DNA Polymerase I