Distribution of G4 motifs and sites of high DNA polymerase and DNA damage across the S. pombe genome. (A) Gray lines above each chromosome indicate the locations of the 446 G4 motifs in the assembled S. pombe genome. Three data tracks are shown below each chromosome: (1) G4 motifs associated with high Pfh1 occupancy (Pfh1, black); (2) G4 motifs associated with high Cdc20 (yellow), γ-H2A (blue) or both (red) in WT cells; and (3) same information as in (2) but for Pfh1-depleted cells. Note that the genome assembly does not include telomeric repeats and has only three complete copies of the approximately 300 rDNA repeats that are located near both telomeres on chromosome III. (B) The location of G4 motifs within a representative rDNA repeat from the left arm of chromosome III (III: 6027–16927). Each of the five G4 motifs (top black bars scaled by G4 motif length) in the 10.9 kb rDNA repeat are on the non-transcribed strand. The arrows denote direction of transcription of the rDNA genes. The locations of Pfh1, Cdc20 and γ-H2A sites are shown below the rDNA annotation track using the same color scheme as in (A). Ter1-3 are natural replication fork barriers, and ARS3001 is a replication origin. (C) Pfh1-associated peaks were validated by ChIP-qPCR using an anti-Myc antibody. Association was calculated as immunoprecipitated DNA divided by input DNA (IP/input). Data present the average of three independent biological replicates and error bars are standard deviations. Three GC-rich sites, three G4 motifs and one tRNA gene (tRNA glu.05) were tested. The tRNA gene was used as a positive control for Pfh1 association. All tested sites were significantly bound by Pfh1 compared to the untagged control strain by two-tailed student t-test P ≤0.01. ChIP, chromatin immunoprecipitation; qPCR, quantitative PCR.