Division of labor of Y-family polymerases in translesion-DNA synthesis for distinct types of DNA damage

Living organisms are continuously under threat from a vast array of DNA-damaging agents, which impact genome DNA. DNA replication machinery stalls at damaged template DNA. The stalled replication fork is restarted via bypass replication by translesion DNA-synthesis polymerases, including the Y-family polymerases Pol eta, Pol iota, and Pol kappa, which possess the ability to incorporate nucleotides opposite the damaged template. To investigate the division of labor among these polymerases in vivo, we generated POL eta(-/-), POL iota(-/-), POL kappa(-/-), double knockout (KO), and triple knockout (TKO) mutants in all combinations from human TK6 cells. TKO cells exhibited a hypersensitivity to ultraviolet (UV), cisplatin (CDDP), and methyl methanesulfonate (MMS), confirming the pivotal role played by these polymerases in bypass replication of damaged template DNA. POL eta(-/-) cells, but not POL iota(-/-) or POL kappa(-/-) cells, showed a strong sensitivity to UV and CDDP, while TKO cells showed a slightly higher sensitivity to UV and CDDP than did POL eta(-/-) cells. On the other hand, TKO cells, but not all single KO cells, exhibited a significantly higher sensitivity to MMS than did wild-type cells. Consistently, DNA-fiber assay revealed that Pol eta plays a crucial role in bypassing lesions caused by UV-mimetic agent 4-nitroquinoline-1-oxide and CDDP, while all three polymerases play complementary roles in bypassing MMS-induced damage. Our findings indicate that the three Y-family polymerases play distinctly different roles in bypass replication, according to the type of DNA damage generated on the template strand.

Automated summary

The study reveals that Y-family polymerases play distinct roles in bypass replication of damaged DNA, exhibiting different sensitivities to various DNA-damaging agents and complementary functions in bypassing specific types of DNA damage.