DNA base excision repair (BER) is a ubiquitous and indispensable defense mechanism against cell death, mutagenesis, and carcinogenesis caused by spontaneously formed DNA damage, such as oxidation and alkylation. Apurinic/apyrimidinic endonuclease (APE1 in mammals) arguably determines the BER rate in cells and thus plays a central role in the repair pathway. Recent findings indicate that APE1 is essential for cellular viability, and that endogenous DNA damage is lethal if left unrepaired. Independently from its repair activity, APE1 interacts with histone deacetylase 1 (HDAC1) to suppress expression of many genes, including the parathyroid hormone gene. APE1 also regulates cJun/cFos and other crucial transcription factors. A high of APE1 is often in are expression observed malignant tumor cells, which more resistant to radio- and chemotherapeutic treatments, suggesting that APE1's repair function increases the chance of survival for cancer cells as well. Thus, APE1 may be either beneficial or harmful to humans, depending on the type of cells in which it is expressed. Understanding the mechanism for regulation of APE1 activity is therefore crucial for evaluating BER efficiency and its consequences in normal and cancer cells. Indeed, cancer predisposition by APE1 deficiency is known and p53 is an upstream regulator of APE1. We have discovered that APE1 is ubiquitinated by mouse double minute 2 (MDM2), and that the ubiquitination is induced by genotoxic stress in a p53-dependent manner. Ubiquitination occurs at specific Lys residues in the APE1 polypeptide, and appears to have significant effects on the functions of APE1 and other cellular proteins, including XRCC1 (X-ray cross-complementation 1), p53, and HDAC1, that interact directly with APE1. The central hypothesis of this project is that APE1 ubiquitination, regulated by MDM2 and p53, modulates the APE1 level and functions in DNA damage response. In order to elucidate the cellular defense mechanism involving this remarkably multifunctional protein, we will: (1) determine the effect of APE1 ubiquitination on its enzymatic and DNA binding activities;(2) unravel interaction of ubiquitinated APE1 with factors in BER and p53 signaling network;and (3) establish the regulation of APE1 ubiquitination in vivo. These parameters are crucial to predict susceptibility of cancer cells against therapeutic drugs that generates genomic instability. With the research facilities provided by Louisiana Cancer Research Consortium (LCRC), this project in the long-term should provide insights for developing new modalities for adjuvant cancer therapies and organ preservation, and for improving the management of other age-related pathophysiology.

Public Health Relevance

DNA is continuously damaged by environmental toxicants like smoke, and by reactive oxygen radicals generated inside of our bodies. Damaged DNA causes mutations and may ultimately lead to cancer and age- related diseases. This project will help us understand how to enhance natural healing of DNA damage, and will help to improve the current methodology for non-surgical cancer treatments, including radiotherapy and chemotherapy.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA098664-09
Application #
8258256
Study Section
Cancer Etiology Study Section (CE)
Program Officer
Okano, Paul
Project Start
2003-03-10
Project End
2014-05-31
Budget Start
2013-06-01
Budget End
2014-05-31
Support Year
9
Fiscal Year
2013
Total Cost
$230,806
Indirect Cost
$75,381
Name
University of Kentucky
Department
Pharmacology
Type
Schools of Medicine
DUNS #
939017877
City
Lexington
State
KY
Country
United States
Zip Code
40506
Busso, C S; Iwakuma, T; Izumi, T (2009) Ubiquitination of mammalian AP endonuclease (APE1) regulated by the p53-MDM2 signaling pathway. Oncogene 28:1616-25
Mantha, Anil K; Oezguen, Numan; Bhakat, Kishor K et al. (2008) Unusual role of a cysteine residue in substrate binding and activity of human AP-endonuclease 1. J Mol Biol 379:28-37
Oezguen, Numan; Schein, Catherine H; Peddi, Srinivasa R et al. (2007) A "moving metal mechanism" for substrate cleavage by the DNA repair endonuclease APE-1. Proteins 68:313-23
Mitra, Sankar; Izumi, Tadahide; Boldogh, Istvan et al. (2007) Intracellular trafficking and regulation of mammalian AP-endonuclease 1 (APE1), an essential DNA repair protein. DNA Repair (Amst) 6:461-9
Chattopadhyay, Ranajoy; Wiederhold, Lee; Szczesny, Bartosz et al. (2006) Identification and characterization of mitochondrial abasic (AP)-endonuclease in mammalian cells. Nucleic Acids Res 34:2067-76
Peddi, Srinivasa R; Chattopadhyay, Ranajoy; Naidu, C V et al. (2006) The human apurinic/apyrimidinic endonuclease-1 suppresses activation of poly(adp-ribose) polymerase-1 induced by DNA single strand breaks. Toxicology 224:44-55
Jackson, Elias B; Theriot, Corey A; Chattopadhyay, Ranajoy et al. (2005) Analysis of nuclear transport signals in the human apurinic/apyrimidinic endonuclease (APE1/Ref1). Nucleic Acids Res 33:3303-12
Izumi, Tadahide; Brown, David B; Naidu, C V et al. (2005) Two essential but distinct functions of the mammalian abasic endonuclease. Proc Natl Acad Sci U S A 102:5739-43
Izumi, Tadahide; Schein, Catherine H; Oezguen, Numan et al. (2004) Effects of backbone contacts 3' to the abasic site on the cleavage and the product binding by human apurinic/apyrimidinic endonuclease (APE1). Biochemistry 43:684-9
Izumi, Tadahide; Wiederhold, Lee R; Roy, Gargi et al. (2003) Mammalian DNA base excision repair proteins: their interactions and role in repair of oxidative DNA damage. Toxicology 193:43-65