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Genes to Cells (2006) 11, 731-744. doi:10.1111/j.1365-2443.2006.00974.x
© 2006 Blackwell Publishing or its licensors
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A human DNA polymerase {eta} complex containing Rad18, Rad6 and Rev1; proteomic analysis and targeting of the complex to the chromatin-bound fraction of cells undergoing replication fork arrest

Mayumi S. Yuasa1,2, Chikahide Masutani1, Akihiko Hirano1, Martin A. Cohn3, Masaru Yamaizumi4, Yoshihiro Nakatani3 and Fumio Hanaoka1,5,*

1 Graduate School of Frontier Biosciences, Osaka University, and SORST, Japan Science and Technology Agency, 1-3 Yamada-Oka, Suita, Osaka 565-0871, Japan
2 Graduate School of Medicine, Osaka University, 2-2 Yamada-Oka, Suita, Osaka 565-0871, Japan
3 Dana-Farber Cancer Institute and Harvard Medical School, Boston, Massachusetts 02115 USA
4 Institute of Molecular Embryology and Genetics, Kumamoto University, Kumamoto 862-0976, Japan
5 Celluler Physiology Laboratory, RIKEN Discovery Research Institute, Wako-shi, Saitama 351-0198, Japan

DNA polymerase eta (Pol{eta}) 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 Pol{eta} leads to xeroderma pigmentosum variant syndrome, clearly indicating that Pol{eta} plays a vital role in preventing skin cancer caused by exposure to sunlight. To further examine Pol{eta} functions and the mechanisms that regulate this important protein, Pol{eta} complexes were purified from HeLa cells over-expressing epitope-tagged Pol{eta}, and polypeptides associated with Pol{eta}, 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 Pol{eta}, Rad18/Rad6 and Rev1, consistent with the requirement for effective TLS by Pol{eta} at sites of DNA lesions.

Communicated by: Hiroyuki Araki

* Correspondence: E-mail: fhanaoka{at}fbs.osaka-u.ac.jp




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