Fig. 4

HydEn-seq-derived evidence for DNA polymerase δ participation in leading-strand synthesis at yeast origins. Red, green, and blue denote polymerases α, δ, and ε, respectively, or DNA tracts synthesized by same. Canonical Okazaki fragments (yellow), synthesized by Pols α (dark), and δ (light) are approximately positioned exemplars. S. cerevisiae origins (identified by Smith et al.²⁸) are oriented such that ARS consensus sequences (ACS) are 5′–3′ beginning at position 0. a Diamonds represent the fraction of DNA strand synthesized by DNA Pols α, δ, and ε (5 bp bins; calculated from rescaled and background subtracted HydEn-seq end densities in pol1-L868M rnh201Δ, pol3-L612M rnh201Δ, and pol2-M644G rnh201Δ strains; see Supplementary Methods). Data for both strands were averaged (opposite strand reflected around +45 bp, the axis of strand symmetry; gray dashed line). Solid curves are regression models. b The fraction of inter-polymerase transfer events outside of canonical Okazaki fragment synthesis (extracted from regression models). The mode of each curve (vertical black line) suggests the most frequent synthesis tract (colored bars above). c–i Schematics of two non-exclusive models of polymerase action at yeast replication origins. DNA strands (colored bars) have the same horizontal scale as in a and b; polymerases (ellipses) and CMG helicases (gray polygons) are exaggerated; other components are omitted. εN and εC indicate N-terminal catalytic and C-terminal CMG-binding Pol ε domains. c Head-to-head dsDNA-binding CMG helicases. d Helicases transition to ssDNA-binding and translocate past one another, N-termini facing directions of travel (gray arrows). e, f Model 1. e Pol α associated with each replisome primes the leading strand that will be synthesized by the other (0.1% probability per bp translocated, from regression). Pol δ extends each nascent-leading strand. f Pols δ collide with respective replisomes, releasing 3′-termini to Pols ε. They assuming synthesis conformation to extend the leading strand. g, h Model 2. g Unidentified Pols α prime leading-strand replication (*; extended by Pols ε). The Pol α associated with each replisome primes the first Okazaki fragment (extended by Pol δ; destined to be longer than average). h Pol δ displaces the 5′ primer terminus of the nascent-leading strand, allowing nick translation or flap excision. i Synthesis patterns from both pathways indicate apparent Pol δ synthesis of both nascent strands at the origin