Selection of template initiation sites and the lengths of RNA primers synthesized by DNA primase are strongly affected by its organization in a multiprotein DNA polymerase alpha complex

Nucleic Acids Res. 1986 Sep 25;14(18):7305-23. doi: 10.1093/nar/14.18.7305.


Synthesis of (p)ppRNA-DNA chains by purified HeLa cell DNA primase-DNA polymerase alpha (pol alpha-primase) was compared with those synthesized by a multiprotein form of DNA polymerase alpha (pol alpha 2) using unique single-stranded DNA templates containing the origin of replication for simian virus 40 (SV40) DNA. The nucleotide locations of 33 initiation sites were identified by mapping G*pppN-RNA-DNA chains and identifying their 5'-terminal ribonucleotide. Pol alpha 2 strongly preferred initiation sites that began with ATP rather than GTP, thus frequently preferring different initiation sites than pol alpha-primase, depending on the template examined. The initiation sites selected in vitro, however, did not correspond to the sites used during SV40 DNA replication in vivo. Pol alpha 2 had the greatest effect on RNA primer size, typically synthesizing primers 1-5 nucleotides long, while pol alpha-primase synthesized primers 6-8 nucleotides long. These differences were observed even at individual initiation sites. Thus, the multiprotein form of DNA primase-DNA polymerase alpha affects selection of initiation sites, the frequency at which the sites are chosen, and length of RNA primers.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Base Sequence
  • DNA Polymerase II / metabolism*
  • DNA Primase
  • DNA Replication
  • DNA, Viral / genetics
  • HeLa Cells / enzymology
  • Humans
  • Multienzyme Complexes / metabolism*
  • Proteins / metabolism*
  • RNA / genetics
  • RNA Nucleotidyltransferases / metabolism*
  • Simian virus 40 / genetics
  • Templates, Genetic
  • Transcription, Genetic


  • DNA, Viral
  • Multienzyme Complexes
  • Proteins
  • RNA
  • DNA Primase
  • RNA Nucleotidyltransferases
  • DNA Polymerase II