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RecBCD-dependent post-UV replication restart in Escherichia coli triggers fork triplication.

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Bibliographic Details
Title: RecBCD-dependent post-UV replication restart in Escherichia coli triggers fork triplication.
Authors: Kouzminovaa, Elena A.1, Cronan, Glen E.1, Kuzminov, Andrei1 kuzminov@illinois.edu
Source: Proceedings of the National Academy of Sciences of the United States of America. 1/27/2026, Vol. 123 Issue 4, p1-12. 34p.
Subjects: Replication fork, DNA replication, Ultraviolet radiation, DNA repair, Escherichia coli, Homologous recombination
Abstract: Ultraviolet (UV) irradiation induces pyrimidine dimers in DNA, which block replication, but are efficiently removed by excision, allowing replication restart. Interestingly, soon after restart, the cells become capable of inducible stable DNA replication (iSDR) without new origin-initiations--suggesting unusual replication topology. The original idea was that the preexisting forks remain dormant even after UV-lesion removal, and when the new origin-initiated forks arrive, they rear-end the dormant forks, creating potential for iSDR. While there is origin-overinitiation after UV, the current idea is that preexisting forks are restarted--making the iSDR potential unclear. We used replication profiling of post-UV cells to investigate reactivation of UV-blocked forks. To compare profile time-sequences, we developed indexation procedure generating nested profile sets. Wild type nested set shows: 1) no replication during the first 30 min post-UV; 2) overinitiation from the origin combined with restart of preexisting forks during the next 30 min post-UV. The restart required RecBCD enzyme to repair double-strand breaks, confirming that the UV-stalled forks disintegrate and need recombinational repair to restart. Preexisting fork restart is separate from new origin-initiations by DnaA, as the dnaA(Ts) mutant at 42 °C restarts preexisting forks normally. Remarkably, the absence of powerful replication wave from origin-initiations in the dnaA mutant reveals a restart-initiated replication wave toward the origin, indicating that the restart mechanism causes fork triplication--potentially explaining the SDR phenomenon. There is no post-UV replication in the dnaA recBC double mutant, consistent with only two pathways of replication resumption in post-UV cells: restart or new origin-initiation. [ABSTRACT FROM AUTHOR]
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Database: Engineering Source
Description
Abstract:Ultraviolet (UV) irradiation induces pyrimidine dimers in DNA, which block replication, but are efficiently removed by excision, allowing replication restart. Interestingly, soon after restart, the cells become capable of inducible stable DNA replication (iSDR) without new origin-initiations--suggesting unusual replication topology. The original idea was that the preexisting forks remain dormant even after UV-lesion removal, and when the new origin-initiated forks arrive, they rear-end the dormant forks, creating potential for iSDR. While there is origin-overinitiation after UV, the current idea is that preexisting forks are restarted--making the iSDR potential unclear. We used replication profiling of post-UV cells to investigate reactivation of UV-blocked forks. To compare profile time-sequences, we developed indexation procedure generating nested profile sets. Wild type nested set shows: 1) no replication during the first 30 min post-UV; 2) overinitiation from the origin combined with restart of preexisting forks during the next 30 min post-UV. The restart required RecBCD enzyme to repair double-strand breaks, confirming that the UV-stalled forks disintegrate and need recombinational repair to restart. Preexisting fork restart is separate from new origin-initiations by DnaA, as the dnaA(Ts) mutant at 42 °C restarts preexisting forks normally. Remarkably, the absence of powerful replication wave from origin-initiations in the dnaA mutant reveals a restart-initiated replication wave toward the origin, indicating that the restart mechanism causes fork triplication--potentially explaining the SDR phenomenon. There is no post-UV replication in the dnaA recBC double mutant, consistent with only two pathways of replication resumption in post-UV cells: restart or new origin-initiation. [ABSTRACT FROM AUTHOR]
ISSN:00278424
DOI:10.1073/pnas.2521115123