ATM and DNA-PK Function Redundantly to Phosphorylate H2AX after Exposure to Ionizing Radiation

By T. Stiff, M. O’Driscoll, N. Rief, Kuniyoshi Iwabuchi, M. Löbrich, P. Jeggo. Cancer Research, 2004

1,057 citations recorded by the source corpus.

H2AX phosphorylation is an early step in the response to DNA damage. It is widely accepted that ATM (ataxia telangiectasia mutated protein) phosphorylates H2AX in response to DNA double-strand breaks (DSBs). Whether DNA-dependent protein kinase (DNA-PK) plays any role in this response is unclear. Here, we show that H2AX phosphorylation after exposure to ionizing radiation (IR) occurs to similar extents in human fibroblasts and in mouse embryo fibroblasts lacking either DNA-PK or ATM but is ablated in ATM-deficient cells treated with LY294002, a drug that specifically inhibits DNA-PK. Additionally, we show that inactivation of both DNA-PK and ATM is required to ablate IR-induced H2AX phosphorylation in chicken cells. We confirm that H2AX phosphorylation induced by DSBs in nonreplicating cells is ATR (ataxia telangiectasia and Rad3-related protein) independent. Taken together, we conclude that under most normal growth conditions, IR-induced H2AX phosphorylation can be carried out by ATM and DNA-PK in a redundant, overlapping manner. In contrast, DNA-PK cannot phosphorylate other proteins involved in the checkpoint response, including chromatin-associated Rad17. However, by phosphorylating H2AX, DNA-PK can contribute to the presence of the damage response proteins MDC1 and 53BP1 at the site of the DSB.

Subfields: Cancer Research, Molecular Biology, Oncology

View Paper: https://www.semanticscholar.org/paper/9163025a7b1912a8d4bb522e33ab5fb9b6e2ab02

How researchers see this paper

The researchers whose trusted networks rank it highest, strongest advocate first. Every researcher gets a different order.

In Cancer Research

1. Stephen P. Jackson
2. M. Lieber
3. F. Alt
4. S. Polo
5. Yosef Shiloh
6. P. Jeggo
7. N. Curtin
8. M. Löbrich

In Molecular Biology

1. Stephen P. Jackson
2. M. Lieber
3. P. Jeggo
4. Yosef Shiloh
5. F. Alt
6. S. Polo
7. G. Iliakis
8. Wenqi Wu

In Oncology

1. M. Lieber
2. Yosef Shiloh
3. Stephen P. Jackson
4. P. Jeggo
5. M. Löbrich
6. S. Polo
7. F. Alt
8. N. Curtin

Citations in the corpus

• An improved zinc-finger nuclease architecture for highly specific genome editing (2007, result): “To confirm the 53BP1 results, we repeated these studies using antibodies for an alternative damage marker, phosphorylated histone H2AX (γH2AX), which is generated in response to DNA damage and forms foci at double-strand break”
• Dynamics of DNA damage response proteins at DNA breaks: a focus on protein modifications. (2011, background): “More generally, it appears that there is a conserved mechanism for apical DDR kinase recruitment to DNA damage sites that involves similar interaction motifs in the C termini of human Ku80-, NBS1-, and ATRIP-mediating interactions with DNA-PKcs, ATM, and ATR, respectively (Falck et al. 2005).”
• ATM signaling facilitates repair of DNA double-strand breaks associated with heterochromatin. (2008, background): “In the absence of ATM, γH2AX foci are less intense (Stiff et al., 2004); this reduced ‘quality’ (but not quantity) of γH2AX foci manifests as a reduction in epitope for IP.”
• Class-switch recombination: interplay of transcription, DNA deamination and DNA repair (2004, background): “It is to be noted, however, that H2AX and 53BP1 are activated in the DSB response by ATM and other phosphatidylinositol 3-kinase (PI3K)-related kinases, such as ATM-related protein (ATR) and DNA-PKc”
• Identification and Characterization of a Novel and Specific Inhibitor of the Ataxia-Telangiectasia Mutated Kinase ATM (2004, background): “This most probably reflects the fact that this site is also targeted by DNA-PK after ionizing radiation (46).”
• The RIDDLE syndrome protein mediates a ubiquitin-dependent signaling cascade at sites of DNA damage. (2009, positive): “ATM promotes the relocalization of many repair and signaling proteins at DNA breaks largely via the phosphorylation of the histone H2A variant H2AX on Ser139 (Rogakou et al., 1998; Stiff et al., 2004).”
• γ-H2AX in recognition and signaling of DNA double-strand breaks in the context of chromatin (2008, background): “DNAPKcs on the other hand, which become activated through its interaction with Ku, after Ku has directly bound to the DNA ends, is likely to have a reduced phosphorylation range requiring longer times for H2AX phosphorylation at longer distances (38,39).”
• The γ‐H2A.X: Is it just a surrogate marker of double‐strand breaks or much more? (2008, neutral): “X [Burma et al., 2001; Stiff et al., 2004].”