Abstract
When a replicative DNA polymerase stalls upon encountering a lesion on the template strand, it is relieved by other low-processivity polymerase(s), which insert nucleotide(s) opposite the lesion, extend by a few nucleotides, and dissociate from the 3′-OH. The replicative polymerase then resumes DNA synthesis. This process, termed translesion replication (TLS) or replicative bypass, may involve at least five different polymerases in mammals, although the participating polymerases and their roles have not been entirely characterized. Using siRNAs originally designed and an alkaline sucrose density gradient sedimentation technique, we verified the involvement of several polymerases in ultraviolet (UV) light-induced TLS in HeLa cells. First, siRNAs to Rev3 or Rev7 largely abolished UV-TLS, suggesting that these 2 gene products, which comprise Polζ, play a main role in mutagenic TLS. Second, Rev1-targeted siRNA also abrogated UV-TLS, indicating that Rev1 is also indispensable to mutagenic TLS. Third, Polη-targeted siRNA also prevented TLS to a greater extent than our expectations. Forth, although siRNA to Polι had no detectable effect, that to Polκ delayed UV-TLS. To our knowledge, this is the first study reporting apparent evidence for the participation of Polκ in UV-TLS.
Highlights
Multiple systems have evolved to manage the genomic photoproducts generated by harmful UV light
We recently reported that caffeine or proteasome inhibitors inhibit UV-translesion replication (TLS) in human cancer cells [36], and that, similar to xeroderma pigmentosum variant (XP-V) cells, UV-TLS was much slower than in normal cells
In Rev3 siRNA-transfected cells, the products remain in smaller size, as depicted by a thin line with open marks, demonstrating these siRNAs prevent UV-TLS (Figure 1(b))
Summary
Multiple systems have evolved to manage the genomic photoproducts generated by harmful UV light. One such system is nucleotide excision repair (NER), which eliminates photoproducts from DNA strands by dual incision on both sides of a damaged base. When a replicative DNA polymerase stalls upon encountering a residual photoproduct on the template strand, it is relieved by other low-processivity polymerase(s), which incorporate nucleotide(s) opposite the lesion, extend by a few nucleotides and dissociate from the 3 -OH. This process, termed translesion replication (TLS) or replicative bypass (reviewed in [1]), is one of the subtle systems that have evolved for the management of genomic photoproducts
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