Abstract
DNA polymerase η (Polη) implements translesion DNA synthesis but has low fidelity in replication. We have previously shown that Polη plays an important role in the genesis of nontargeted mutations at undamaged DNA sites in cells exposed to the carcinogen N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). Here, we report that MNNG-induced Polη expression in an interferon regulatory factor 1 (IRF1)-dependent manner in human cells. Mutagenesis analysis showed that four critical residues (Arg-82, Cys-83, Asn-86, and Ser-87) located in the IRF family conserved DNA binding domain-helix α3 were involved in DNA binding and POLH transactivation by IRF1. Furthermore, Polη up-regulation induced by IRF1 was responsible for the increase of mutation frequency in a SupF shuttle plasmid replicated in the MNNG-exposed cells. Interestingly, IRF1 was acetylated by the histone acetyltransferase CBP in these cells. Lys → Arg substitution revealed that Lys-78 of helix α3 was the major acetylation site, and the IRF1-K78R mutation partially inhibited DNA binding and its transcriptional activity. Thus, we propose that IRF1 activation is responsible for MNNG-induced Polη up-regulation, which contributes to mutagenesis and ultimately carcinogenesis in cells.
Highlights
Pol implements translesion DNA synthesis but has low fidelity in replication
We found that overexpressed wild-type interferon regulatory factor 1 (IRF1) significantly increased the mutation frequency of the supF tRNA gene in the plasmid replicated in FL cells, the K78R mutant induced fewer mutations, whereas the DNA-binding Domain (DBD) 4-mutant did not increase the mutation frequency over control (Table 1)
We demonstrated that exposure to the carcinogen MNNG increased human POLH gene expression through IRF1 transactivation in human FL cells
Summary
Pol implements translesion DNA synthesis but has low fidelity in replication. Results: Acetylation-stabilized IRF1 transactivates the POLH gene in response to the chemical carcinogen N-methyl-NЈ-nitroN-nitrosoguanidine (MNNG). IRF1 Transactivated POLH Responsible for MNNG-induced Mutations undamaged DNA sequences (nontargeted mutations) in mammalian cells, and this is caused by a decrease of DNA replication fidelity resulting from aberrant expression of TLS DNA polymerases upon MNNG exposure [13, 14]. Using genomic and proteomic methodologies, we further revealed that many genes and proteins are involved in the response to MNNG exposure, including those participating in the regulation of transcription, metabolism, cytoskeletal organization, the cell cycle, cell proliferation, signal transduction, and transport (20 –22) These findings help our understanding of the molecular mechanisms of MNNG-caused damage, how chemical carcinogens regulate the low-fidelity TLS DNA polymerases that contribute to mutagenesis and carcinogenesis remains unclear. We propose that IRF1-dependent transcriptional regulation of TLS DNA polymerases is probably a new mutagenic and carcinogenic pathway stimulated by environmental chemical carcinogens
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