Quantitative sensing and signalling of single-stranded DNA during the DNA damage response

The DNA damage checkpoint senses the presence of DNA lesions and controls the cellular response thereto. A crucial DNA damage signal is single-stranded DNA (ssDNA), which is frequently found at sites of DNA damage and recruits the sensor checkpoint kinase Mec1-Ddc2. However, how this signal - and therefore the cell's DNA damage load - is quantified, is poorly understood. Here, we use genetic manipulation of DNA end resection to induce quantitatively different ssDNA signals at a site-specific double strand break in budding yeast and identify two distinct signalling circuits within the checkpoint. The local checkpoint signalling circuit leading to γH2A phosphorylation is unresponsive to increased amounts of ssDNA, while the global checkpoint signalling circuit, which triggers Rad53 activation, integrates the ssDNA signal quantitatively. The global checkpoint signal critically depends on the 9-1-1 and its downstream acting signalling axis, suggesting that ssDNA quantification depends on at least two sensor complexes.