DNA polymerase eta (Polη) is responsible for efficient translesion synthesis (TLS) past cis‐syn cyclobutane thymine dimers (TT dimers), the major DNA lesions induced by UV irradiation. Loss of human Polη leads to xeroderma pigmentosum variant syndrome, clearly indicating that Polη plays a vital role in preventing skin cancer caused by exposure to sunlight. To further examine Polη functions and the mechanisms that regulate this important protein, Polη complexes were purified from HeLa cells over‐expressing epitope‐tagged Polη, and polypeptides associated with Polη, including Rad18, Rad6 and Rev1, were identified by a combination of mass spectrometry and Western blot analysis. The chromatin‐bound fractions of cells subjected to UV irradiation, S phase synchronization, or S phase arrest were specifically enriched in such complexes. These results suggest that arrested replication forks strengthen interactions among Polη, Rad18/Rad6 and Rev1, consistent with the requirement for effective TLS by Polη at sites of DNA lesions.