Fig. 4: DDK regulates long-range resection via phosphorylation of Dna2.
a Serine 236 of Dna2 is phosphorylated in a DDK-dependent manner. Data from phospho-proteomic experiment of Fig. 1d. Heat-map depicts the z-score. See also Supplementary Fig. 4a, b. b Scheme of Dna2 phosphorylation sites. Green: CDK target sites previously identified48; Orange: DDK target site S236, within S/T-S/T motif. c Serine 236 phosphorylation contributes to DDK-dependent Dna2 phosphorylation shift in vivo. Cells of the indicated strains were arrested in G1 or M-phase and samples collected and loaded on a gel to monitor the Dna2 phospho-shift. Data are representative of n = 3 biological replicates. See also Supplementary Fig. 4c, d. d dna2-S236A strain is sensitive to CPT, when EXO1 is deleted. Five-fold serial dilutions of WT, exo1Δ, dna2-S236A, and dna2-S236A exo1Δ strains were grown on YPD plates or YPD plates supplemented with CPT at the indicated concentrations. Data are representative of n = 3 biological replicates. e Depletion of Dbf4 induces defects in STR-Dna2-mediated long-range resection. ssDNA accumulation upon DSB induction was measured via qPCR after digestion with the RsaI restriction nuclease at sites with indicated distances from the DSB. n = 3 biological replicates, shown is mean with values of replicates, error bars denote SD. Reported p-values were calculated using a two-tailed unpaired t-test. See also Supplementary Fig. 4f, g. f DDK-dependent resection defect accumulates in the absence of EXO1. Resection was measured as in (e), but in a background where resection was carried out by Sae2-MRX and STR-Dna2. n = 3 biological replicates, shown is mean with values of replicates, error bars denote SD. Reported p-values were calculated using a two-tailed unpaired t-test. See also Supplementary Fig. 4f, h. Source data are provided as a Source Data file.
