Veronica G. Godoy

A single amino acid governs enhanced activity of DinB DNA polymerases on damaged templates

Translesion synthesis (TLS) by Y-family DNA polymerases is a chief mechanism of DNA damage tolerance. Such TLS can be accurate or error-prone, as it is for bypass of a cyclobutane pyrimidine dimer by DNA polymerase η (XP-V or Rad30) or bypass of a (6-4) TT photoproduct by DNA polymerase V (UmuD′2C), respectively. Although DinB is the only Y-family DNA polymerase conserved among all domains of life, the biological rationale for this striking conservation has remained enigmatic. Here we report that the Escherichia coli dinB gene is required for resistance to some DNA-damaging agents that form adducts at the N2-position of deoxyguanosine (dG). We show that DinB (DNA polymerase IV) catalyses accurate TLS over one such N2-dG adduct (N2-furfuryl-dG), and that DinB and its mammalian orthologue, DNA polymerase κ, insert deoxycytidine (dC) opposite N2-furfuryl-dG with 10–15-fold greater catalytic proficiency than opposite undamaged dG. We also show that mutating a single amino acid, the ‘steric gate’ residue of DinB (Phe13 → Val) and that of its archaeal homologue Dbh (Phe12 → Ala), separates the abilities of these enzymes to perform TLS over N2-dG adducts from their abilities to replicate an undamaged template. We propose that DinB and its orthologues are specialized to catalyse relatively accurate TLS over some N2-dG adducts that are ubiquitous in nature, that lesion bypass occurs more efficiently than synthesis on undamaged DNA, and that this specificity may be achieved at least in part through a lesion-induced conformational change.

Y‐family DNA polymerases respond to DNA damage‐independent inhibition of replication fork progression

In Escherichia coli, the Y‐family DNA polymerases Pol IV (DinB) and Pol V (UmuD2′C) enhance cell survival upon DNA damage by bypassing replication‐blocking DNA lesions. We report a unique function for these polymerases when DNA replication fork progression is arrested not by exogenous DNA damage, but with hydroxyurea (HU), thereby inhibiting ribonucleotide reductase, and bringing about damage‐independent DNA replication stalling. Remarkably, the umuC122∷Tn5 allele of umuC, dinB, and certain forms of umuD gene products endow E. coli with the ability to withstand HU treatment (HUR). The catalytic activities of the UmuC122 and DinB proteins are both required for HUR. Moreover, the lethality brought about by such stalled replication forks in the wild‐type derivatives appears to proceed through the toxin/antitoxin pairs mazEF and relBE. This novel function reveals a role for Y‐family polymerases in enhancing cell survival under conditions of nucleotide starvation, in addition to their established functions in response to DNA damage.

Transcriptional Modulator NusA Interacts with Translesion DNA Polymerases in Escherichia coli

NusA, a modulator of RNA polymerase, interacts with the DNA polymerase DinB. An increased level of expression of dinB or umuDC suppresses the temperature sensitivity of the nusA11 strain, requiring the catalytic activities of these proteins. We propose that NusA recruits translesion DNA synthesis (TLS) polymerases to RNA polymerases stalled at gaps, coupling TLS to transcription.

Proficient and Accurate Bypass of Persistent DNA Lesions by DinB DNA Polymerases

Despite nearly universal conservation through evolution, the precise function of the DinB/pol κ branch of the Y-family of DNA polymerases has remained unclear. Recent results suggest that DinB orthologs from all domains of life proficiently bypass replication blocking lesions that may be recalcitrant to DNA repair mechanisms. Like other translesion DNA polymerases, the error frequency of DinB and its orthologs is higher than the DNA polymerases that replicate the majority of the genome. However, recent results suggest that some Y-family polymerases, including DinB and pol κ, bypass certain types of DNA damage with greater proficiency than an undamaged template. Moreover, they do so relatively accurately. The ability to employ this mechanism to manage DNA damage may be especially important for types of DNA modification that elude repair mechanisms. For these lesions, translesion synthesis may represent a more important line of defense than for other types of DNA damage that are more easily dealt with by other more accurate mechanisms.