The Many Roles of PCNA in Eukaryotic DNA Replication

Abstract

Proliferating cell nuclear antigen (PCNA) plays critical roles in many aspects of DNA replication and replication-associated processes, including translesion synthesis, error-free damage bypass, break-induced replication, mismatch repair, and chromatin assembly. Since its discovery, our view of PCNA has evolved from a replication accessory factor to the hub protein in a large protein–protein interaction network that organizes and orchestrates many of the key events at the replication fork. We begin this review article with an overview of the structure and function of PCNA. We discuss the ways its many interacting partners bind and how these interactions are regulated by posttranslational modifications such as ubiquitylation and sumoylation. We then explore the many roles of PCNA in normal DNA replication and in replication-coupled DNA damage tolerance and repair processes. We conclude by considering how PCNA can interact physically with so many binding partners to carry out its numerous roles. We propose that there is a large, dynamic network of linked PCNA molecules at and around the replication fork. This network would serve to increase the local concentration of all the proteins necessary for DNA replication and replication-associated processes and to regulate their various activities.

Introduction

Since its discovery in the late 1970s, our view of proliferating cell nuclear antigen (PCNA) and its roles in DNA replication and genome maintenance has expanded considerably. PCNA was originally identified as the target of an autoimmune antibody derived from patients with systemic lupus erythematosus [1]. This protein was later shown to be one produced predominantly in proliferating and transformed cells [2], [3], [4]. By the middle of the 1980s, the involvement of PCNA in DNA replication was suggested based on its pattern of staining throughout the cell cycle [5].

Definitive evidence of a role for PCNA in DNA replication came a couple years later with the discovery that PCNA is required for the replication of simian virus 40 in vitro [6], [7]. It was soon realized that PCNA was an auxiliary protein for DNA polymerase delta (pol δ) that increases its activity by making it more processive [8], [9], [10]. PCNA was subsequently shown to be an auxiliary factor for DNA polymerase epsilon (pol ɛ) [11], [12], [13], [14]. By the early 1990s, the role of PCNA came to be viewed as being the processivity factor of eukaryotic replicative polymerases.

An understanding of how PCNA confers high processivity to DNA polymerases was achieved when the X-ray crystal structure of PCNA was determined [15]. PCNA was shown to be a ring-shaped trimer similar to the structure of the Escherichia coli beta clamp determined a couple years earlier [16]. By the middle of the 1990s, it was known that PCNA is loaded onto double-stranded DNA by replication factor C (RFC) [17], [18], where the PCNA functions as a sliding clamp that binds and anchors polymerases onto the DNA.

As more and more PCNA-interacting partners were identified, it became clear that PCNA is not simply a processivity factor for replicative polymerases. It interacts with and regulates the activities of many proteins involved in Okazaki fragment maturation [19], [20], mismatch repair [21], nucleotide excision repair [22], and translesion synthesis [23], [24], [25], [26]. It also interacts with proteins involved in other processes such as cell cycle control [27], [28], [29], sister chromatid cohesion [30], epigenetic inheritance [31], and S-phase-specific proteolysis [32]. By the early 2000s, PCNA came to be viewed as an important hub protein that is critical for organizing and orchestrating events at the replication fork and other sites of DNA synthesis.

Since the early 2000s, it has become clear that the regulation of several DNA metabolic processes is governed by posttranslational modifications of PCNA, most notably ubiquitylation and sumoylation [33], [34]. Ubiquitylation of PCNA promotes translesion synthesis via the recruitment of translesion synthesis polymerases to stalled replication forks [35]. Sumoylation of PCNA inhibits recombination via the recruitment of antirecombinases to sites of DNA synthesis [36], [37].

In this chapter, we will describe the many roles of PCNA in eukaryotic DNA replication and in replication-associated processes. We will begin by discussing the features of the structure and function of PCNA common to all of its roles. Then we will focus on its roles in normal DNA replication, translesion synthesis and error-free damage bypass, break-induced replication, mismatch repair, and replication-coupled nucleosome assembly.