Escherichia Coli DNA Polymerase IV (Pol IV), but Not Pol II, Dynamically Switches with a Stalled Pol III* Replicase

Overview

Journal J Bacteriol

Publisher American Society for Microbiology

Specialty Microbiology

Date 2012 May 1

PMID 22544274

Citations 31

Authors

Justin M H Heltzel

Robert W Maul

David W Wolff

Mark D Sutton

Affiliations

Soon will be listed here.

Abstract

The dnaN159 allele encodes a temperature-sensitive mutant form of the β sliding clamp (β159). SOS-induced levels of DNA polymerase IV (Pol IV) confer UV sensitivity upon the dnaN159 strain, while levels of Pol IV ∼4-fold higher than those induced by the SOS response severely impede its growth. Here, we used mutations in Pol IV that disrupted specific interactions with the β clamp to test our hypothesis that these phenotypes were the result of Pol IV gaining inappropriate access to the replication fork via a Pol III*-Pol IV switch relying on both the rim and cleft of the clamp. Our results clearly demonstrate that Pol IV relied on both the clamp rim and cleft interactions for these phenotypes. In contrast to the case for Pol IV, elevated levels of the other Pols, including Pol II, which was expressed at levels ∼8-fold higher than the normal SOS-induced levels, failed to impede growth of the dnaN159 strain. These findings suggest that the mechanism used by Pol IV to switch with Pol III* is distinct from those used by the other Pols. Results of experiments utilizing purified components to reconstitute the Pol III*-Pol II switch in vitro indicated that Pol II switched equally well with both a stalled and an actively replicating Pol III* in a manner that was independent of the rim contact required by Pol IV. These results provide compelling support for the Pol III*-Pol IV two-step switch model and demonstrate important mechanistic differences in how Pol IV and Pol II switch with Pol III*.

Citing Articles

Thumb-domain dynamics modulate the functional repertoire of DNA-Polymerase IV (DinB).

Okeke D, Lidman J, Matecko-Burmann I, Burmann B Nucleic Acids Res. 2023; 51(13):7036-7052.

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Observing protein dynamics during DNA-lesion bypass by the replisome.

Wilkinson E, Spenkelink L, van Oijen A Front Mol Biosci. 2022; 9:968424.

PMID: 36213113 PMC: 9534484. DOI: 10.3389/fmolb.2022.968424.

Compartmentalization of the replication fork by single-stranded DNA-binding protein regulates translesion synthesis.

Chang S, Thrall E, Laureti L, Piatt S, Pages V, Loparo J Nat Struct Mol Biol. 2022; 29(9):932-941.

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Direct visualization of translesion DNA synthesis polymerase IV at the replisome.

Tuan P, Gilhooly N, Marians K, Kowalczykowski S Proc Natl Acad Sci U S A. 2022; 119(39):e2208390119.

PMID: 36122225 PMC: 9522359. DOI: 10.1073/pnas.2208390119.

During Translesion Synthesis, Escherichia coli DinB89 (T120P) Alters Interactions of DinB (Pol IV) with Pol III Subunit Assemblies and SSB, but Not with the β Clamp.

Scotland M, Homiski C, Sutton M J Bacteriol. 2022; 204(4):e0061121.

PMID: 35285726 PMC: 9017331. DOI: 10.1128/jb.00611-21.

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