Replisome structure and conformational dynamics underlie fork progression past obstacles

Abstract

Replisomes are multiprotein complexes that unzip the parental helix and duplicate the separated strands during genome replication. The antiparallel structure of DNA poses unique geometric constraints to the process, and the replisome has evolved unique dynamic features that solve this problem. Interestingly, the solution to duplex DNA replication has been co-opted to solve many other important problems that replisomes must contend with during the duplication of long chromosomes. For example, along its path the replisome will encounter lesions and DNA-bound proteins. Recent studies show that the replisome can circumvent lesions on either strand, using the strategy normally applied to the lagging strand synthesis. Circumventing lesions can also be assisted by other proteins that transiently become a part of the replisome. The replisome must also contend with DNA-binding proteins and recent studies reveal a fascinating process that enables it to bypass RNA polymerase without stopping.

Figure 1

Replisome structure at the E. coli replication fork. (a) DnaB helicase encircles the lagging strand, and primase targets DnaB for RNA primer synthesis. The ssDNA is coated with SSB (not shown). Two Pol III molecules are connected through contact with C-terminal extensions of the two τ subunits of the central clamp loader, which also bind DnaB. The two Pol III core molecules each interact with a β clamp for processive synthesis. Owing to the antiparallel duplex, the connection of the two polymerases results in forming a ‘trombone loop’ during lagging strand synthesis. (b) Components of replisomes in different replication systems.

Figure 2

Structures of the bacterial polymerase and the β sliding clamp. (a) The Pol III α subunit from E. coli (pdb code, 2HNH), reproduced with permission from [14]. (b) The E. coli β clamp on primed DNA (pdb code, 3BEP), reproduced with permission from [19 ••]. (c) Model of T. aquaticus Pol III α subunit bound to DNA (pdb code, 3e0d), docked onto the E. coli β-DNA complex (pdb code, 3BEP), along with E. coli Pol IV attached to β (pdb code, 1UNN). Reproduced with permission from [21 ••].

Figure 3

Polymerase recycling paths on the lagging strand. The lagging strand polymerase undergoes ‘collision release’ upon completing an Okazaki fragment, leaving the clamp on DNA (top path), and undergoes ‘premature release’ when it is signaled to dissociate from the clamp before completing an Okazaki fragment (bottom path).

Figure 4

The replisome circumvents blocks on the leading strand. (a) Pol III stalls at a template lesion and hops to a new clamp on an RNA primer synthesized by primase. (b) A TLS polymerase switches with Pol III on the clamp and extends DNA over the template lesion, producing a mutation. (c) Codirectional encounters of the replisome with RNA polymerase result in the dissociation of the RNA polymerase and recruitment of the mRNA to continue synthesis.