Large-scale unbiased phosphoproteomics analysis of Tel1-deleted yeast reveal dozens of novel Tel1-dependent phosphorylation events. The canonical S/T-Q motif is enriched amongst these Tel1-dependent sites, but a novel D/E-S/T motif is even more highly enriched. Phosphoproteomics of possible downstream acidophilic kinases Dun1, Rad53, and Hrr25 are performed to determine if these sites may be indirectly Tel1-dependent, but the D/E-S/T sites are not dependent on any of them.