Abstract
DNA constantly undergoes chemical modification due to endogenous and exogenous mutagens. The DNA base excision repair (BER) pathway is the frontline mechanism handling the majority of these lesions, and primarily involves a DNA incision and subsequent resealing step. It is imperative that these processes are extremely well-coordinated as unrepaired DNA single strand breaks (SSBs) can be converted to DNA double strand breaks during replication thus triggering genomic instability. However, the mechanism(s) governing the BER process are poorly understood. Here we show that accumulation of unrepaired SSBs triggers a p53/Sp1-dependent downregulation of APE1, the endonuclease responsible for the DNA incision during BER. Importantly, we demonstrate that impaired p53 function, a characteristic of many cancers, leads to a failure of the BER coordination mechanism, overexpression of APE1, accumulation of DNA strand breaks and results in genomic instability. Our data provide evidence for a previously unrecognized mechanism for coordination of BER by p53, and its dysfunction in p53-inactivated cells.
Highlights
Genomic DNA is inherently unstable due to its intrinsic chemical nature
In order to be able to assess if unrepaired single strand breaks (SSBs) feed back to apyrimidinic endonuclease 1 (APE1), we forced TIG-1 normal human fibroblasts to accumulate endogenously generated SSBs by creating an artificial base excision repair (BER) imbalance through transient knockdown of X-ray repair crosscomplementing protein 1 (XRCC1)
BER is a fundamental housekeeping DNA repair system that deals with the majority of endogenously generated DNA lesions
Summary
Genomic DNA is inherently unstable due to its intrinsic chemical nature. It is estimated that as many as 10 000 DNA lesions/cell/day can arise under physiological conditions [1]. Accumulation of these lesions results in mutations and leads to genomic instability, which is a hallmark of cancer cells [2]. The DNA base excision repair (BER) pathway is a frontline mechanism preventing genomic instability, as it contributes to cell defence against most endogenous and exogenous sources of genotoxic lesions. BER is responsible for the elimination of base alterations (e.g. oxidation, alkylation) and DNA single strand breaks (SSBs); the latter can arise either spontaneously, or as a consequence of BER processing of damaged DNA bases [3]. The resulting abasic site (AP-site) is processed by an apurinic/apyrimidinic endonuclease, which cleaves the phosphodiester bond 5 to the baseless site, generating a SSB. In mammalian cells apurinic/apyrimidinic endonuclease 1 (APE1) accounts for the majority of the AP-site cleavage activity [4,5] and is a crucial enzyme for mammalian DNA repair. In canonical BER, the resulting SSB is eventually sealed by a protein complex containing DNA polymerase , X-ray repair crosscomplementing protein 1 (XRCC1) and DNA ligase III␣ [3], where XRCC1 acts as a scaffold protein to coordinate the formation and the stability of the DNA polymerase -XRCC1–DNA ligase III␣ complex on SSBs [7]
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