AUF1 recognizes 8-oxo-guanosine embedded in DNA and stimulates APE1 endoribonuclease activity.

Abstract

The existence of modified rNMPs embedded in genomic DNA, as a consequence of oxidative stress conditions, including 8-oxo-guanosine and ribose monophosphate abasic site (rAP), has been recently highlighted by several works including ours. Although the activity of APE1, a key enzyme of the base excision repair pathway, in the repair of rAP sites results as efficient as the canonical deoxyribose monophosphate abasic sites (dAP), its incision repair activity on 8-oxo-guanosine is very weak. The aims of this work were to: i) identify proteins able to specifically bind 8-oxo-guanosine embedded in DNA and promote APE1 endoribonuclease on this lesion and ii) characterize the molecular and biological relevance of this interaction using human cancer cell lines. By using an unbiased proteomic approach, we discovered that AUF1 actively recognizes 8-oxo-guanosine and stimulates the APE1 enzymatic activity on this DNA lesion. By using orthogonal approaches, we found that: i) the interaction between AUF1 and APE1 is modulated by H2O2-treatment; ii) depletion of APE1 and AUF1 causes the accumulation of single- and double- strand breaks; iii) both proteins are involved in modulating the formation of DNA:RNA hybrids. These data establish unexpected functions of AUF1 in modulating genome stability, and improve our knowledge of APE1 biology with respect to 8-oxo-guanosine embedded in DNA. By showing a novel function of AUF1, our findings shed new light on the process of maintenance of genome stability in mammalian cells towards oxidative stress-related damages.