TY - JOUR
T1 - 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
AU - Vishwanatha, Jamboor K.
AU - Yamaguchi, Masamitsu
AU - Depamphilis, Melvin L.
AU - Baril, Earl F.
N1 - Funding Information:
ACKNOWLEDGMENTS This research was supported by National Institutes of Health Grants P30 12708, CA 15187, and CA 15579 fron the National Canoer Institute and American Cancer Society Grant MV114. J.K.V. is a Fellow of the Leukemia Sooiety of America. We are grateful to Lois Eager and Sandra Johnson for their assistance in preparation of this manuscript.
PY - 1986/9/11
Y1 - 1986/9/11
N2 - Synthesis of (p)ppRNA-DNA chains by purified HeLa cell DNA primase-DNA polymerase α (pol α-primase) was compared with those synthesized by a multiprotein form of DNA polymerase α (pol α2) 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 ribo-nucleotide. Pol α2 strongly preferred initiation sites that began with ATP rather than GTP, thus frequently preferring different initiation sites than pol α-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 vitro. Pol α2 had the greatest effect on RNA primer size, typically synthesizing primers 1-5 nucleotides long, while pol α-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 a affects selection of initiation sites, the frequency at which the sites are chosen, and length of RNA primers.
AB - Synthesis of (p)ppRNA-DNA chains by purified HeLa cell DNA primase-DNA polymerase α (pol α-primase) was compared with those synthesized by a multiprotein form of DNA polymerase α (pol α2) 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 ribo-nucleotide. Pol α2 strongly preferred initiation sites that began with ATP rather than GTP, thus frequently preferring different initiation sites than pol α-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 vitro. Pol α2 had the greatest effect on RNA primer size, typically synthesizing primers 1-5 nucleotides long, while pol α-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 a affects selection of initiation sites, the frequency at which the sites are chosen, and length of RNA primers.
UR - http://www.scopus.com/inward/record.url?scp=0023056980&partnerID=8YFLogxK
U2 - 10.1093/nar/14.18.7305
DO - 10.1093/nar/14.18.7305
M3 - Article
C2 - 2429260
AN - SCOPUS:0023056980
SN - 0305-1048
VL - 14
SP - 7305
EP - 7323
JO - Nucleic Acids Research
JF - Nucleic Acids Research
IS - 18
ER -