DNA Replication: Mammalian

Abstract

Deoxyribonucleic acid (DNA) replication is an evolutionarily semi‐conserved process that involves a mechanism to unwind the parent DNA strand, the synthesis of identical daughter strands, and steps to terminate the sequence. Owing to the massive amount of genetic information needed to be copied error‐free, the replication of DNA within the mammalian chromosome is highly complex, requiring the simultaneous firing of multiple origins and processing of DNA at defined sites within the cells. This action requires the strict regulation and precise timing of numerous proteins, enzymes and substrates. These components critical to DNA replication will associate within these compartments in the cell to form replication factories during processing. Although many of the basic functions of DNA synthesis are well‐defined, there is still much yet to be understood regarding the machinery involved in the initiation, synthesis and termination of mammalian DNA replication.

Key Concepts:

  • Mammalian DNA replication occurs at specific regions within the cell.

  • Proteins necessary for DNA replication come together to form ‘replication factories’ within the nucleus.

  • The manner and timing of mammalian DNA replication is largely determined in G1 of the cell cycle.

  • The selection and timing of each replication site is controlled by a combination of primary sequences, kinase activation, transcription, and local chromatin environment.

  • The proteins thus far identified at the mammalian cell DNA replication fork include the MCM helicase complex, DNA pol α‐primase, PCNA, RFC, DNA pol δ, FEN1, RPA, DNA ligase I, topoisomerase I and II and RNAse H.

  • Termination of mammalian DNA replication is thought to be dependent on physical barriers within the replication factories, as opposed to actual termination sequences within the DNA structure.

  • In the event of replication fork stalling, repair pathways must be activated to halt cell cycle progression and repair the damage.

Keywords: DNA replication; mammalian cells; replication factories; initiation; replication fork; termination; replication fork stalling

Figure 1.

The ORC (origin recognition complex) binds double‐stranded DNA upon initiation. This signals Cdc6 interaction, resulting in helicase loading (Mcm2–7) as a double hexamer. In order to stabilise the complex, Cdc45 and GINS bind to Mcm2–7 (CMG) in order to unwind the DNA.

Figure 2.

Leading and lagging strand synthesis of mammalian DNA at the replication fork. After unwinding of the parental DNA, replication protein A (RPA) binds, stabilizing the DNA pol α‐primase complex. Following primer synthesis, replicating factor C (RFC) loads proliferating cell nuclear antigen (PCNA) onto the leading strand. PCNA then acts as a scaffold to load polymerase δ (pol δ), continuing synthesis in the 5′→3′ direction. On the lagging strand, pol α‐primase creates Okazaki fragments, which are extended by pol δ. When these fragments converge, a single‐stranded flap is formed. This flap is then cleaved by flap endonuclease 1 (FEN1) and ribonucleic acidase (RNAse H). The resulting nick is sealed by DNA ligase 1.

Figure 3.

A model of ssDNA damage response. (1) DNA damage (red “D”) in the leading or lagging strand stalls DNA polymerase. As DNA helicases continue to unwind DNA, RPA covers accumulating ssDNA. (2) ATRIP responds by binding to RPA, signalling ATR binding. (3) One example of the DNA damage sensor is the Rad17‐RFC complex loading the 9‐1‐1 clamp (Rad9‐Rad1‐Hus1) to DNA in order to activate ATR through mediator proteins (4). (5) Activated ATR can phosphorylate effector proteins such as Chk1.

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Smith, Shanna J, Li, Caroline M, Hickey, Robert J, and Malkas, Linda H(Nov 2014) DNA Replication: Mammalian. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0001041.pub3]