Structure-Function Studies Of DNA Replication Fidelity

工作概述：该项目的长期目标是(1)了解从病毒到人类等生物体中控制细胞存活和基因组稳定性的DNA复制和修复过程的保真度和效率，以及(2)了解环境压力如何干扰这些过程，从而导致细胞毒性、诱变和对人类健康的不利影响。今年，在了解几种不同DNA聚合酶的功能方面取得了进展，包括线粒体pol γ和T7 DNA聚合酶（A家族），核复制pol δ （B家族），DNA修复pol λ和mu （X家族），以及翻译合成pol iota （Y家族）。这些研究揭示了细胞准确复制正常DNA以及被环境因子破坏的DNA所使用的策略，以避免突变和细胞毒性以及随之而来的不利健康后果。本项目已与Z01-ES-65046结合。

Summary of Work: The long-term goals of this project are (1) to understand the fidelity and efficiency of the DNA replication and repair processes that control cell survival and genome stability in organisms from viruses to humans, and (2) to understand how environmental stress perturbs these processes, resulting in cytotoxicity, mutagenesis and adverse effects on human health. This year, progress was made towards understanding the functions of several different DNA polymerases, including mitochondrial pol gamma and T7 DNA polymerase (family A), nuclear replicative pol delta (family B), DNA repair pols lambda and mu (family X), and translesion synthesis pol iota (family Y). These studies are revealing the strategies used by cells to accurately replicate normal DNA as well as DNA damaged by environmental agents, in a manner that avoids mutagenesis and cytotoxicity and the accompanying adverse health consequences. This project has been combined with Z01-ES-65046. Several repair processes operate prior to replication to remove DNA damage generated by endogenous cellular metabolism or exposure to environmental stress. These repair processes provide undamaged substrates for efficient and highly accurate DNA replication. High replication fidelity depends on the ability of DNA polymerases to select correct rather than incorrect nucleotides for incorporation into DNA without adding or deleting nucleotides. This selectivity is a prime determinant of fidelity at the replication fork and also during DNA synthesis associated with repair. Exonucleolytic proofreading of mismatches can further increase DNA replication fidelity. When DNA damage is not repaired prior to replication, certain lesions can stall progression of the replication fork. Under these circumstances, cell cycle checkpoints, translesion synthesis reactions and recombinational repair processes operate to maintain genome stability and improve cell survival. This project investigates these replication fidelity processes by studying the relationships between the structures of A, B, X, Y and RT family DNA polymerases and their accessory proteins, their biochemical properties and their putative functions in vivo. A dual approach is used involving in vitro biochemical studies with in vivo genetic studies in yeast and mice and humans.