New research from a team of genome scientists and DNA damage response (DDR) experts breaks new ground in understanding the function of a protein currently limited in clinical trials for cancer treatments. The new investigaton shows how ATM-mediated signaling is induced by DNA single-strand breaks (SSBs) for DNA damage repair – illuminating the distinct mechanisms of SSB-induced ATM kinase and shedding an important light on APE1 function. In the new study, researchers demonstrate their use of plasmid-based SSB structures to examine APE1’s critical role in DNA damage response signaling pathways. APE1 or apurynic endonuclease/redox factor Ref-1, is an ubiquitous and vital protein that acts as an essential master regulator of this response, highly contributing to the maintenance of the genome stability.
APE1 is a dual function protein involved both in the base excision repair (BER) pathways of DNA lesions, acting as the major apurinic/apyrimidinic endonuclease, and in eukaryotic transcriptional regulation of gene expression. This effect is obtained as a redox co-activator of different transcription factors such as Egr-1, nuclear factor-κB (NF-κB), tumor suppressor p53, hypoxia inducible factor-1α (HIF-1α), CREB, AP-1 and paired box-containing proteins (Pax) in different cell systems. The vital role of APE1 seems to be due to its fundamental activity in the base excision repair pathway of DNA lesions. A more recent finding has identified APE1’s role in mediating production of single-strand DNA (ssDNA) breaks in gene promoters during repair of targeted base oxidation lesions caused by oxygen radicals generated during physiologic signaling.
Thus, defects in the APE1-mediated step in BER pathway could be linked to altered gene expression besides altering transcription factor state. Among the other findings, the publication details results showing SSB induces ATM activation prior to ATR, temporarily arresting cell cycle progression as DNA attempts to undergo repair. And – central to the study’s significance – the team discovered direct evidence for the active role played by APE1 in single strand break-induced ATM DDR signaling. Scientists demonstrated that APE1 promotes SSB-induced ATM DDR through at least two mechanisms: APE1 exonuclease activity-mediated SSB processing and APE1-mediated direct recruitment of ATM to SSBs. These knowlegde may contribute to future therapeutic inhibitor options for a range of human diseases, such as cancer and heart failure.
DNA damage response is an evolutionary pathway, designed to maintain genome integrity as human cells are under near-constant attack, by both internal and external factors. DNA lesions generally trigger DDR – signaling a “cascade,” as the study puts it, of transcription activation, repair, and cell cycle arrests. While DNA damage repair research is far more established for cellular response to double-strand breaks (DSBs), scientists say knowledge gaps surrounding SSBs have persisted. In particular, molecular biologists have had little to no direct evidence about the precise function of APE1 at SSB sites. In the past, researchers have generally regarded functional DSB studies to be more urgent, as this type of DNA damage is closely associated with cancer. However, SSBs can foreshadow the development of more lethal double-strand damage.
In addition, SSB damage occurs far more frequently, more than 10,000 times per day in a human cell. In each instance, repair mechanisms must take place to protect genome integrity. The accumulation of SSBs over time is thought to contribute to or cause cancer and disease like chronic heart failure (CHF). Intense oxidative stress develops in the heart with CHF, with depletion of inner antioxidants like GSH and Trx-1. Reactive oxygen species (ROS), moreover, lead to the activation of ATM kinase itself and mitochondia impairment, again, amplify and sustain the process leading to telomere shortening and the onset of multiple SSBs and /or DSBs, until myocardial cells undergo programmed cells death (apoptosis). Further, this brings forth an intriguing inquiry on whether APE1 over-expression can be shown consistently to activate ATM in the absence of DNA.
Previous studies have elucidated several molecular mechanisms of how the ATM-dependent DDR is activated. ATM phosphorylation is rapidly induced by DSBs in trans, and has been widely utilized as an indicator of ATM activation. Furthermore, histone acetyltransferase Tip60 and its cofactor TRRAP promote ATM activation at DSB sites via ATM interaction and acetylation. In addition, oxidative stressor hydrogen peroxide can directly activate ATM through distinct ATM homodimerization. In this context, APE1 is also a redox protein and might well modulate this process. Taken together, study data have uncovered mechanisms of ATM activation by SSBs and identified APE1 as a direct ATM activator. On the oncology field, this could be taken advantage of by using APE and ATM inhibitors which may synergize in cancer cell killing with alkylating chemogrugs.
On the clinical cardiology side, heart failure cellular loss could be prevented by redox conditioning of the APE1-ATM interaction. Handling CHF with antioxidant drugs would actually act with a double mechanism, one of which would be the direct scavenging of ROS before they could generate SSBs or DSBs. In this case prevention of SSB accumulation or persistent DDR activation may become a new therapeutic strategy against heart failure.
- Edited by Dr. Gianfrancesco Cormaci, PhD, specialist in Clinical Biochemisty.
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