A novel role for Gadd45 alpha in base excision repair: Modulation of APE1 activity by the direct interaction of Gadd45 alpha with PCNA

Abstract

The growth arrest and DNA damage inducible, alpha (Gadd45 alpha) protein regulates DNA repair by interacting with proliferating cell nuclear antigen (PCNA). Our previous study suggested a potential role for Gadd45 alpha in the base excision repair (BER) pathway by affecting apurinic/apyrimidinic endonuclease 1 (APE1) protein in addition to its accepted role in nucleotide excision repair (NER). Here, we investigated whether the interaction of Gadd45 alpha with PCNA affects APE1 activity. To address this issue, we used a siRNA directed to Gadd45 alpha and a form of Gadd45 alpha with a mutation to the predicted site of PCNA binding. There was a reduction of APE1 activity in cells transfected with the Gadd45 alpha siRNA. Furthermore, the interaction of Gadd45 alpha with PCNA and APE1 was lower in cells transfected with mutant Gadd45 alpha compared with cells transfected with wild-type Gadd45 alpha. Indeed, we observed that the APE1 activity in the Gadd45 alpha-interacting complex was significantly lower in cells that overexpress mutant Gadd45 alpha, compared with cells that overexpress wild-type Gadd45 alpha. We conclude that the PCNA binding site on Gadd45 alpha plays a critical role in modulating the interaction with PCNA and APE1, affecting BER activity. These results provide novel insights into the mechanisms by which BER activity is modulated, although the interaction of Gadd45 alpha with APE1 needs to be clarified. (c) 2013 Elsevier Inc. All rights reserved.