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Post-replicative base excision repair in replication foci.

M Otterlei¹, E Warbrick, T A Nagelhus

¹Institute of Cancer Research and Molecular Biology, Faculty of Medicine, Norwegian University of Science and Technology, N-7005 Trondheim, Norway.

The EMBO Journal

|July 7, 1999

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The major nuclear uracil-DNA glycosylase (UNG2) removes uracil during DNA replication. This base excision repair (BER) process involves proliferating cell nuclear antigen (PCNA) and replication protein A (RPA) at the replication fork.

Area of Science:

Molecular Biology
DNA Repair Mechanisms
Cellular Biochemistry

Background:

Base excision repair (BER) is a critical pathway for maintaining genomic stability.
BER can be completed via alternative routes, some involving DNA replication proteins.
Uracil incorporation into DNA can arise from deamination of cytosine or misincorporation of dUTP.

Purpose of the Study:

To investigate the role of uracil-DNA glycosylase (UNG2) in DNA replication and repair.
To determine the interaction of UNG2 with key replication factors like PCNA and RPA.
To elucidate the formation of a base excision repair complex at the replication fork.

Main Methods:

Monitoring UNG2 expression and localization during S phase using BrdUrd incorporation.

Related Experiment Videos

Assessing uracil removal in isolated nuclei using neutralizing antibodies against UNG2.

Investigating molecular interactions between UNG2, PCNA, and RPA via two-hybrid assays, peptide SPOT assays, and ELISAs.

Main Results:

UNG2 levels increase during S phase and co-localize with replication foci.
UNG2 is responsible for the rapid removal of uracil from newly synthesized DNA.
Direct molecular interactions were confirmed between UNG2, PCNA, and RPA.

Conclusions:

UNG2 plays a significant role in the rapid post-replicative removal of incorporated uracil.
A functional base excision repair complex comprising UNG2, RPA, and PCNA forms near the replication fork.
This BER complex likely contributes to maintaining genome integrity during DNA replication.