Repair of U/G and U/A in DNA by UNG2-associated repair complexes takes place predominantly by short-patch repair both in proliferating and growth-arrested cells

TY - JOUR

T1 - Repair of U/G and U/A in DNA by UNG2-associated repair complexes takes place predominantly by short-patch repair both in proliferating and growth-arrested cells

AU - Akbari, Mansour

AU - Otterlei, Marit

AU - Pena Diaz, Javier

AU - Aas, Per Arne

AU - Kavli, Bodil

AU - Liabakk, Nina B

AU - Hagen, Lars

AU - Imai, Kohsuke

AU - Durandy, Anne

AU - Slupphaug, Geir

AU - Krokan, Hans E

PY - 2004

Y1 - 2004

N2 - Nuclear uracil-DNA glycosylase UNG2 has an established role in repair of U/A pairs resulting from misincorporation of dUMP during replication. In antigen-stimulated B-lymphocytes UNG2 removes uracil from U/G mispairs as part of somatic hypermutation and class switch recombination processes. Using antibodies specific for the N-terminal non-catalytic domain of UNG2, we isolated UNG2-associated repair complexes (UNG2-ARC) that carry out short-patch and long-patch base excision repair (BER). These complexes contain proteins required for both types of BER, including UNG2, APE1, POLbeta, POLdelta, XRCC1, PCNA and DNA ligase, the latter detected as activity. Short-patch repair was the predominant mechanism both in extracts and UNG2-ARC from proliferating and less BER-proficient growth-arrested cells. Repair of U/G mispairs and U/A pairs was completely inhibited by neutralizing UNG-antibodies, but whereas added recombinant SMUG1 could partially restore repair of U/G mispairs, it was unable to restore repair of U/A pairs in UNG2-ARC. Neutralizing antibodies to APE1 and POLbeta, and depletion of XRCC1 strongly reduced short-patch BER, and a fraction of long-patch repair was POLbeta dependent. In conclusion, UNG2 is present in preassembled complexes proficient in BER. Furthermore, UNG2 is the major enzyme initiating BER of deaminated cytosine (U/G), and possibly the sole enzyme initiating BER of misincorporated uracil (U/A).

AB - Nuclear uracil-DNA glycosylase UNG2 has an established role in repair of U/A pairs resulting from misincorporation of dUMP during replication. In antigen-stimulated B-lymphocytes UNG2 removes uracil from U/G mispairs as part of somatic hypermutation and class switch recombination processes. Using antibodies specific for the N-terminal non-catalytic domain of UNG2, we isolated UNG2-associated repair complexes (UNG2-ARC) that carry out short-patch and long-patch base excision repair (BER). These complexes contain proteins required for both types of BER, including UNG2, APE1, POLbeta, POLdelta, XRCC1, PCNA and DNA ligase, the latter detected as activity. Short-patch repair was the predominant mechanism both in extracts and UNG2-ARC from proliferating and less BER-proficient growth-arrested cells. Repair of U/G mispairs and U/A pairs was completely inhibited by neutralizing UNG-antibodies, but whereas added recombinant SMUG1 could partially restore repair of U/G mispairs, it was unable to restore repair of U/A pairs in UNG2-ARC. Neutralizing antibodies to APE1 and POLbeta, and depletion of XRCC1 strongly reduced short-patch BER, and a fraction of long-patch repair was POLbeta dependent. In conclusion, UNG2 is present in preassembled complexes proficient in BER. Furthermore, UNG2 is the major enzyme initiating BER of deaminated cytosine (U/G), and possibly the sole enzyme initiating BER of misincorporated uracil (U/A).

KW - Cell Division

KW - Cell Line

KW - DNA

KW - DNA Glycosylases

KW - DNA Repair

KW - DNA-(Apurinic or Apyrimidinic Site) Lyase

KW - DNA-Binding Proteins

KW - HeLa Cells

KW - Humans

KW - Macromolecular Substances

KW - Precipitin Tests

KW - Substrate Specificity

KW - Uracil

KW - Uracil-DNA Glycosidase

U2 - 10.1093/nar/gkh872

DO - 10.1093/nar/gkh872

M3 - Journal article

C2 - 15479784

SN - 0305-1048

VL - 32

SP - 5486

EP - 5498

JO - Nucleic Acids Research

JF - Nucleic Acids Research

IS - 18

ER -