Abstract
High fidelity (HiFi) DNA polymerases (Pols) perform the bulk of DNA synthesis required to duplicate genomes in all forms of life. Their structural features, enzymatic mechanisms, and inherent properties are well-described over several decades of research. HiFi Pols are so accurate that they become stalled at sites of DNA damage or lesions that are not one of the four canonical DNA bases. Once stalled, the replisome becomes compromised and vulnerable to further DNA damage. One mechanism to relieve stalling is to recruit a translesion synthesis (TLS) Pol to rapidly synthesize over and past the damage. These TLS Pols have good specificities for the lesion but are less accurate when synthesizing opposite undamaged DNA, and so, mechanisms are needed to limit TLS Pol synthesis and recruit back a HiFi Pol to reestablish the replisome. The overall TLS process can be complicated with several cellular Pols, multifaceted protein contacts, and variable nucleotide incorporation kinetics all contributing to several discrete substitution (or template hand-off) steps. In this review, we highlight the mechanistic differences between distributive equilibrium exchange events and concerted contact-dependent switching by DNA Pols for insertion, extension, and resumption of high-fidelity synthesis beyond the lesion.
Highlights
DNA damage can come from a variety of endogenous and exogenous sources and persist within the genome into the synthesis (S) phase of the cell cycle (Hakem, 2008)
All of the examples described in the Postlesion: Resuming High Fidelity Synthesis, a Final Substitution section are mechanisms used to limit synthesis by a translesion synthesis (TLS) Pol after insertion to maintain downstream genome fidelity
TLS Pols are more mutagenic on undamaged templates than on suitable damaged substrates; so strict regulation outside of a specific TLS insertion is necessary
Summary
DNA damage can come from a variety of endogenous and exogenous sources and persist within the genome into the synthesis (S) phase of the cell cycle (Hakem, 2008). The covalent addition of Ub to the back side of PCNAK164 and the flexibility inherent within the unstructured C-terminal tails containing these motifs allow these TLS Pols to adopt several conformations on PCNA in inactive carrier or active polymerizing states (Shen et al, 2021) Based on this multivalency for trimeric mUb-PCNA, there are at least six specific contact points for recruiting and interacting with Pols, providing a landing platform for assemblies (Figure 2) in what has been described previously as a “tool-belt” model (Kath et al, 2014; Boehm et al, 2016b; Cranford et al, 2017). Recruitment of a TLS Pol to a stalled HiFi holoenzyme may be brought in by specific contacts with PCNA (Figure 2) or combined with Pol-Pol interactions creating a transient supraholoenzyme to initiate a switch only to have the HiFi Pol and/or PCNA dissociate in a subsequent exchange for insertion opposite a lesion
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