From bacteria to mammalian cells, damaged DNA is sensed and targeted by DNA repair pathways. A distinguishing feature of eukaryotes is that kinases play central roles in coordination of the DNA damage response. DNA damage signaling kinases were identified over 20 years ago and linked to imposition of the cell cycle checkpoint concept proposed by Weinert and Hartwell. Following their identification, a role for DNA damage signaling kinases in protecting genome integrity was proposed, but evidence for their direct involvement in DNA repair was lacking. As a result, the initial perception was that the role of these kinases in genome maintenance was an indirect result of their cell cycle checkpoint function. As more substrates of DNA damage signaling kinases were identified, it became clear that these kinases play more direct roles in preserving the integrity of the genome. The central theme of my thesis is how one of these kinases, Mec1/ATR, directly suppresses spontaneous gross chromosomal rearrangements and my efforts to find the critical substrates that mediate this function. Phosphoproteomic mass spectrometry has been a key tool for my investigations of Mec1 action. My employment of this technology in tracking the activity of Mec1 kinase has resulted in the largest single phosphoproteomic data set in budding yeast, containing over 30,000 phosphorylation events occurring in roughly 2/3 of the yeast proteome. A secondary theme of my thesis details my efforts to extract biologically meaningful phosphorylation events from the thousands of phosphosites identified by proteomic mass spectrometry.