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Caffeine Abolishes the Ultraviolet-Induced REV3 Translesion Replication Pathway in Mouse Cells
Division of Genetic Biochemistry, The National Institute of Health and Nutrition, Shinjuku-ku, Tokyo 162-8636, Japan
School of Medicine, Tokai University, Isehara-shi, Kanagawa-ken 259-1193, Japan
Present address: Kanagawa Institute of Technology, Atsugi-shi, Kanagawa 243-0292, Japan
* Author to whom correspondence should be addressed.
Received: 16 September 2011; in revised form: 8 November 2011 / Accepted: 17 November 2011 / Published: 29 November 2011
Abstract: When a replicative DNA polymerase stalls upon encountering a photoproduct on the template strand, it is relieved by other low-processivity polymerase(s), which insert nucleotide(s) opposite the lesion. Using an alkaline sucrose density gradient sedimentation technique, we previously classified this process termed UV-induced translesion replication (UV-TLS) into two types. In human cancer cells or xeroderma pigmentosum variant (XP-V) cells, UV-TLS was inhibited by caffeine or proteasome inhibitors. However, in normal human cells, the process was insensitive to these reagents. Reportedly, in yeast or mammalian cells, REV3 protein (a catalytic subunit of DNA polymerase ζ) is predominantly involved in the former type of TLS. Here, we studied UV-TLS in fibroblasts derived from the Rev3-knockout mouse embryo (Rev3KO-MEF). In the wild-type MEF, UV-TLS was slow (similar to that of human cancer cells or XP-V cells), and was abolished by caffeine or MG-262. In 2 cell lines of Rev3KO-MEF (Rev3−/− p53−/−), UV-TLS was not observed. In p53KO-MEF, which is a strict control for Rev3KO-MEF, the UV-TLS response was similar to that of the wild-type. Introduction of the Rev3 expression plasmid into Rev3KO-MEF restored the UV-TLS response in selected stable transformants. In some transformants, viability to UV was the same as that in the wild-type, and the death rate was increased by caffeine. Our findings indicate that REV3 is predominantly involved in UV-TLS in mouse cells, and that the REV3 translesion pathway is suppressed by caffeine or proteasome inhibitors.
Keywords: alkaline sucrose density gradient sedimentation; caffeine; DNA polymerase ζ; proteasome inhibitor; TLS; translesion replication; Rev3 knockout
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Takezawa, J.; Aiba, N.; Kajiwara, K.; Yamada, K. Caffeine Abolishes the Ultraviolet-Induced REV3 Translesion Replication Pathway in Mouse Cells. Int. J. Mol. Sci. 2011, 12, 8513-8529.
Takezawa J, Aiba N, Kajiwara K, Yamada K. Caffeine Abolishes the Ultraviolet-Induced REV3 Translesion Replication Pathway in Mouse Cells. International Journal of Molecular Sciences. 2011; 12(12):8513-8529.
Takezawa, Jun; Aiba, Naomi; Kajiwara, Kagemasa; Yamada, Kouichi. 2011. "Caffeine Abolishes the Ultraviolet-Induced REV3 Translesion Replication Pathway in Mouse Cells." Int. J. Mol. Sci. 12, no. 12: 8513-8529.