Physiological Tolerance to ssDNA Enables Strand Uncoupling during DNA Replication

Abstract

Summary: It has been long assumed that normally leading strand synthesis must proceed coordinated with the lagging strand to prevent strand uncoupling and the pathological accumulation of single-stranded DNA (ssDNA) in the cell, a dogma recently challenged by in vitro studies in prokaryotes. Here, we report that human DNA polymerases can function independently at each strand in vivo and that the resulting strand uncoupling is supported physiologically by a cellular tolerance to ssDNA. Active forks rapidly accumulate ssDNA at the lagging strand when POLA1 is inhibited without triggering a stress response, despite ssDNA formation being considered a hallmark of replication stress. Acute POLA1 inhibition causes a lethal RPA exhaustion, but cells can duplicate their DNA with limited POLA1 activity and exacerbated strand uncoupling as long as RPA molecules suffice to protect the elevated ssDNA. Although robust, this uncoupled mode of DNA replication is also an in-built weakness that can be targeted for cancer treatment. : Using specific POLA1 inhibitors, Ercilla et al. show that DNA synthesis works independently at leading and lagging strands and can be “uncoupled” in vivo. The authors show that cells can naturally tolerate ssDNA formation during DNA replication as long as it is protected by the surplus of RPA molecules. Keywords: DNA replication, polymerase alpha, POLA1, CD437, lagging strand, strand uncoupling, RPA, ssDNA, replication catastrophe, ATR