Distinct roles of DNA polymerases delta and epsilon at the replication fork in Xenopus egg extracts.
DNA polymerases delta and epsilon (Poldelta and Polepsilon) are widely thought to be the major DNA polymerases that function in elongation during DNA replication in eukaryotic cells. However, the precise roles of these polymerases are still unclear. Here we comparatively analysed DNA replication in Xenopus egg extracts in which Poldelta or Polepsilon was immunodepleted. Depletion of either polymerase resulted in a significant decrease in DNA synthesis and accumulation of short nascent DNA products, indicating an elongation defect. Moreover, Poldelta depletion caused a more severe defect in elongation, as shown by sustained accumulation of both short nascent DNA products and single-stranded DNA gaps, and also by elevated chromatin binding of replication proteins that function more frequently during lagging strand synthesis. Therefore, our data strongly suggest the possibilities that Poldelta is essential for lagging strand synthesis and that this function of Poldelta cannot be substituted for by Polepsilon.
Fukui
Tomoyuki
T
Department of Biochemistry and Molecular Biology, Research Institute for Microbial Diseases, Osaka University, 3-1 Yamada-oka, Suita, Osaka 565-0871, Japan.
Yamauchi
Kazumi
K
Muroya
Taketo
T
Akiyama
Masahiro
M
Maki
Hisaji
H
Sugino
Akio
A
Waga
Shou
S
D000818:Animals; D000906:Antibodies / Q000494:pharmacology; D002457:Cell Extracts / Q000032:analysis; D002843:Chromatin / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004257:DNA Polymerase II / Q000502:physiology*; D004258:DNA Polymerase III / Q000502:physiology*; D004261:DNA Replication / Q000187:drug effects; D004268:DNA-Binding Proteins / Q000378:metabolism; D045585:Flap Endonucleases / Q000378:metabolism; D010063:Ovum / Q000737:chemistry / Q000201:enzymology; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism; D011485:Protein Binding; D051716:Replication Protein A; D051818:Replication Protein C; D014981:Xenopus / Q000235:genetics* / Q000378:metabolism
D000906:Antibodies; D002457:Cell Extracts; D002843:Chromatin; D004268:DNA-Binding Proteins; C022262:Okazaki fragments; D018809:Proliferating Cell Nuclear Antigen; D051716:Replication Protein A; D004247:DNA; D004257:DNA Polymerase II; D004258:DNA Polymerase III; D045585:Flap Endonucleases; D051818:Replication Protein C