Requirement of mammalian DNA polymerase-β in base-excision repair

Abstract

Synthesis of DNA by DNA polymerase-beta is distributive on single-stranded DNA templates, but short DNA gaps with a 5' PO4 in the gap are filled processively to completion. In vitro studies have suggested a role of beta-polymerase in different types of DNA repair. However, the significance of these studies to the in vivo role of beta-polymerase has remained unclear. Because genetic studies are essential for determining the physiological role of a gene, we established embryonic fibroblast cell lines homozygous for a deletion mutation in the gene encoding DNA polymerase-beta. Extracts from these cell lines were found to be defective in uracil-initiated base-excision repair. The beta-polymerase-deleted cells are normal in viability and growth characteristics, although they exhibit increased sensitivity to monofunctional DNA-alkylating agents, but not to other DNA-damaging agents. Both the deficiency in base-excision repair and hypersensitivity to DNA-alkylating agents are rescued following stable transfection with a wild-type beta-polymerase minitransgene. These studies demonstrate that beta-polymerase functions specifically in base-excision repair in vivo.

Mammalian base excision repair by DNA polymerases δ and ε

“…Pol b seems to catalyze the excision of the 5'-terminal deoxyribose phosphate more e ciently than basic molecules (Matsumoto and Kim, 1995). This might explain why in vivo Pol b is the DNA polymerase of election for one nucleotide replacement reactions (Sobol et al, 1996).…”

“…Whole cell extracts from Pol b-deleted mouse ®broblasts (Sobol et al, 1996) were fractionated to separate Pol d and Pol e. In a ®rst fractionation step over phosphocellulose, the extract was depleted from PCNA and RP-A. The second fractionation step over hydroxylapatite led to a separation of Pol a from Pol d/e.…”

“…Pol d and/or Pol e have been implicated in the synthesis step of the other two major excision repair processes, nucleotide excision repair (Shivji et al, 1995) and mismatch repair (Longley et al, 1997) which both require PCNA (Shivji et al, 1992, Umar et al, 1996. As far as BER is concerned the characteristics of mouse ®broblasts cell lines with a homozygous deletion of Pol b (Sobol et al, 1996) give the most clear evidence that Pol b functions speci®cally in this pathway in the replacement of a single damaged nucleotide. In contrast, however, the identity of the Pols involved in the long-patch BER is still matter of debate.…”

A unified view of base excision repair: Lesion-dependent protein complexes regulated by post-translational modification

“…This mechanism or process of complex formation appears to provide an increase in specificity and efficiency to the BER pathway, thereby facilitating the maintenance of genome integrity by preventing the accumulation of highly toxic repair intermediates [83]. Decreased concentrations of just one protein within this pathway could significantly alter the balance of complex formation, resulting in reduced repair capacity and an increased exposure to toxic BER intermediates, such as has been observed in pol ß deficiency in mouse cells [79,[91][92][93], in animal models [94] and in human cells [62].…”

Mutator phenotype of BCR – ABL transfected Ba/F3 cell lines and its association with enhanced expression of DNA polymerase β

“…Pol b, structurally the simplest of the ®ve known mammalian DNA polymerases, is believed to function primarily in the repair of damaged bases (Sobol et al, 1996). It is a monomeric protein of 335 amino acids (39 kDa) that lacks exonuclease activities and appears to be the least accurate of the eukaryotic DNA polymerases in vitro (Kunkel, 1985).…”