Publication Synopsis

ATM and ATR substrate analysis reveals extensive protein networks responsive to DNA damage

In the May 25th issue of Science a research team led by Stephen Elledge published a study identifying a broad network of kinases and substrates responsive to DNA damage. Several phosphoproteome assays were employed to identify over 700 targets of DNA damage-activated kinases. An IPA Core analysis of this set of DNA Damage Response (DDR) targets revealed a significant enrichment for proteins involved in DNA replication, recombination and repair. IPA further revealed that these substrates organized into interaction modules involved in DNA replication, DNA repair, and cell cyle processes. Their integrated approach of identifying a DDR-response specific phosphoproteome, followed by functional and network analysis in IPA expands the scope of the DNA damage response far beyond what had been previously identified, and provides a foundation for a systems biology level understanding of cellular response to DNA damage.

Elledge et al combined multiple phosphoprotein detection technologies (anti-phospho Ser-Gln antibodies, peptide IP and SILAC) in order to ensure an unbiased, comprehensive approach to identifying kinase substrates regulated in response to DNA damage. As a result, they identified over 905 phosphorylations sites on 700 proteins that demonstrated an increase in phosphorylation of more than 4-fold after DNA damage was induced by ionizing radiation.

IPA’s Functional Analysis demonstrated that the phosphorylation targets of ionizing radiation were highly enriched for the processes DNA replication, recombination and repair. Significant enrichments were also observed for proteins in the categories of cell cycle, gene expression, and cell signaling as well as an unexpected enrichment for RNA post-translational modification proteins, including splicing factors.

IPA’s Network Analysis further identified interacting modules involved in DNA replication (ORC module, MCM module), and DNA repair (mismatch repair module, excision repair module). Many of the components of these modules had not been previously linked to DNA damage signaling. IPA Canonical Pathway analysis identified an enriched set of interactions between the DNA damage response targets and the AKT-insulin signaling pathway, leading the authors to hypothesize that DDR might induce a survival signal through activation of AKT.

This comprehensive examination of the targets of DNA damage activated kinases and the interaction modules and processes they participate in leads to a better understanding of the significant alterations in cellular physiology induced by the DNA damage response.