Parental histone deposition on the replicated strands promotes error-free DNA damage tolerance and regulates drug resistance

Ctf4 is a conserved replisome component with multiple roles in DNA metabolism. To investigate connections between Ctf4-mediated processes involved in drug resistance, we conducted a suppressor screen of ctf4Δ sensitivity to the methylating agent MMS. We uncovered that mutations in Dpb3 and Dpb4 components of polymerase ε result in the development of drug resistance in ctf4Δ via their histone-binding function. Alleviated sensitivity to MMS of the double mutants was not associated with rescue of ctf4Δ defects in sister chromatid cohesion, replication fork architecture, or template switching, which ensures error-free replication in the presence of genotoxic stress. Strikingly, the improved viability depended on translesion synthesis (TLS) polymerase-mediated mutagenesis, which was drastically increased in ctf4 dpb3 double mutants. Importantly, mutations in Mcm2–Ctf4–Polα and Dpb3–Dpb4 axes of parental (H3–H4)2 deposition on lagging and leading strands invariably resulted in reduced error-free DNA damage tolerance through gap filling by template switch recombination. Overall, we uncovered a chromatin-based drug resistance mechanism in which defects in parental histone transfer after replication fork passage impair error-free recombination bypass and lead to up-regulation of TLS-mediated mutagenesis and drug resistance.

Ctf4是一类保守的复制体（replisome）组分，在DNA代谢中发挥多重功能。为探究Ctf4介导的、与耐药性相关的生物学过程之间的关联，我们针对ctf4缺失突变体（ctf4Δ）对甲基化试剂MMS（methylating agent MMS）的敏感性开展了抑制筛选实验。我们发现，DNA聚合酶ε（polymerase ε）的Dpb3与Dpb4亚基发生突变后，可通过其组蛋白结合功能使ctf4Δ菌株产生耐药性。上述双突变体对MMS的敏感性得以缓解，但这并未挽救ctf4Δ在姐妹染色单体黏连（sister chromatid cohesion）、复制叉结构（replication fork architecture）或模板转换（template switching）方面的缺陷——模板转换可确保遗传毒性应激（genotoxic stress）环境下的无错复制。值得注意的是，菌株存活率的提升依赖于跨损伤合成（translesion synthesis, TLS）聚合酶介导的诱变作用，且ctf4Δ dpb3Δ双突变体中的该诱变过程显著增强。重要的是，在滞后链（lagging strand）与前导链（leading strand）的亲本(H3–H4)2沉积通路中，Mcm2–Ctf4–Polα与Dpb3–Dpb4两条信号轴发生突变后，均会通过模板转换重组（template switch recombination）填补间隙的方式，无一例外地降低了无错DNA损伤耐受能力。综上，我们揭示了一种基于染色质的耐药性机制：复制叉通过后亲本组蛋白转移的缺陷会损害无错重组旁路，并最终导致跨损伤合成介导的诱变作用上调与耐药性产生。