In response to DNA double-strand breaks or oxidative stress, ATM-dependent
DNA damage response (DDR) is activated to maintain genome integrity.
However, it remains elusive whether and how DNA single-strand breaks (SSBs)
activate ATM. Here, we provide direct evidence in Xenopus egg extracts that
ATM-mediated DDR is activated by a defined SSB structure. Our mechanistic
studies reveal that APE1 promotes the SSB-induced ATM DDR through APE1
exonuclease activity and ATM recruitment to SSB sites. APE1 protein can form
oligomers to activate the ATM DDR in Xenopus egg extracts in the absence of
DNA and can directly stimulate ATM kinase activity in vitro. Our findings reveal
distinct mechanisms of the ATM-dependent DDR activation by SSBs in
eukaryotic systems and identify APE1 as a direct activator of ATM kinase.