Successful DNA replication requires carefully regulated mechanisms to overcome numerous obstacles that naturally occur throughout chromosomal DNA. Scattered across the genome are tightly bound proteins, such as transcription factors and nucleosomes, that are necessary for cell function, but that also have the potential to impede timely DNA replication. Using biochemically reconstituted systems, we show that two transcription factors, yeast Reb1 and Tbf1, and a tightly positioned nucleosome, are strong blocks to the strand displacement DNA synthesis activity of DNA polymerase δ. Although the block imparted by Tbf1 can be overcome by the DNA-binding activity of the single-stranded DNA-binding protein RPA, efficient DNA replication through either a Reb1 or a nucleosome block occurs only in the presence of the 5'-3' DNA helicase Pif1. The Pif1-dependent stimulation of DNA synthesis across strong protein barriers may be beneficial during break-induced replication where barriers are expected to pose a problem to efficient DNA bubble migration. However, in the context of lagging strand DNA synthesis, the efficient disruption of a nucleosome barrier by Pif1 could lead to the futile re-replication of newly synthetized DNA. In the presence of FEN1 endonuclease, the major driver of nick translation during lagging strand replication, Pif1-dependent stimulation of DNA synthesis through a nucleosome or Reb1 barrier is prevented. By cleaving the short 5' tails generated during strand displacement, FEN1 eliminates the entry point for Pif1. We propose that this activity would protect the cell from potential DNA re-replication caused by unwarranted Pif1 interference during lagging strand replication.

中文翻译：

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Pif1、RPA 和 FEN1 调节 DNA 聚合酶 δ 在 DNA 合成过程中克服蛋白质障碍的能力。

成功的 DNA 复制需要精心调节的机制来克服在整个染色体 DNA 中自然发生的众多障碍。基因组中散布着紧密结合的蛋白质，例如转录因子和核小体，它们是细胞功能所必需的，但也有可能阻碍 DNA 的及时复制。使用生化重组系统，我们表明两个转录因子，酵母 Reb1 和 Tbf1，以及一个紧密定位的核小体，是 DNA 聚合酶 δ 的链置换 DNA 合成活性的强阻滞剂。尽管可以通过单链 DNA 结合蛋白 RPA 的 DNA 结合活性克服 Tbf1 赋予的阻断，但通过 Reb1 或核小体阻断的有效 DNA 复制仅在存在 5'-3' DNA 的情况下发生解旋酶 Pif1。Pif1 依赖的 DNA 合成刺激跨越强大的蛋白质屏障可能在断裂诱导的复制过程中是有益的，其中屏障预计会对有效的 DNA 气泡迁移构成问题。然而，在滞后链 DNA 合成的背景下，Pif1 对核小体屏障的有效破坏可能导致新合成的 DNA 无效的重新复制。在 FEN1 核酸内切酶（滞后链复制期间切口平移的主要驱动因素）存在的情况下，通过核小体或 Reb1 屏障对 DNA 合成的 Pif1 依赖性刺激被阻止。通过切割链置换过程中产生的 5' 短尾，FEN1 消除了 Pif1 的入口点。