Werner Syndrome (WS) is an inherited disease characterized by premature onset of aging, increased cancer incidence, and genomic instability. The WS gene encodes a 1,432-amino acid polypeptide (WRN) with a central domain homologous to the RecQ family of DNA helicases. Purified WRN unwinds DNA with 3'-->5' polarity, and also possesses 3'-->5' exonuclease activity. Elucidation of the physiologic function(s) of WRN may be aided by the identification of WRN-interacting proteins. We show here that WRN functionally interacts with DNA polymerase delta (pol delta), a eukaryotic polymerase required for DNA replication and DNA repair. WRN increases the rate of nucleotide incorporation by pol delta in the absence of proliferating cell nuclear antigen (PCNA) but does not stimulate the activity of eukaryotic DNA polymerases alpha or epsilon, or a variety of other DNA polymerases. Moreover, we show that functional interaction with WRN is mediated through the third subunit of pol delta: i.e., Pol32p of Saccharomyces cerevisae, corresponding to the recently identified p66 subunit of human pol delta. Absence of the third subunit abrogates stimulation by WRN, and stimulation is restored by reconstituting the three-subunit enzyme. Our findings suggest that WRN may facilitate pol delta-mediated DNA replication and/or DNA repair and that disruption of WRN-pol delta interaction in WS cells may contribute to the previously observed S-phase defects and/or the unusual sensitivity to a limited number of DNA damaging agents.