Replication protein A

Steps in DNA synthesis

Replication Protein A (RPA) is a protein complex crucial in the DNA replication and DNA repair processes of eukaryotes. It is a single-stranded DNA-binding protein (ssDNA-binding protein) that plays a pivotal role in various cellular activities, including DNA replication, DNA repair, recombination, and cell cycle regulation. RPA is highly conserved across eukaryotes, underscoring its essential role in cell biology.

Structure

RPA is a heterotrimeric complex composed of three subunits: RPA1, RPA2, and RPA3, each contributing to the complex's DNA-binding and protein-protein interaction capabilities. The subunits are named based on their molecular weights: RPA1 (70 kDa), RPA2 (32 kDa), and RPA3 (14 kDa). The structure of RPA allows it to bind ssDNA with high affinity and specificity, a critical feature for its function in DNA metabolism.

Function

The primary function of RPA is to stabilize and protect single-stranded DNA (ssDNA) intermediates that form during replication, repair, and recombination processes. By binding to ssDNA, RPA prevents the formation of secondary structures and protects the DNA from nucleolytic degradation. Its roles include:

• DNA Replication: RPA stabilizes the unwound DNA at replication forks, facilitating the activity of DNA polymerases and other replication proteins.
• DNA Repair: In DNA repair pathways, such as nucleotide excision repair (NER) and homologous recombination (HR), RPA binds to ssDNA regions to recruit and stabilize repair proteins at the damage site.
• Recombination: During recombination, RPA binds to ssDNA generated after strand invasion, playing a crucial role in the repair of double-strand breaks and the restart of stalled replication forks.
• Cell Cycle Regulation: RPA interacts with various proteins involved in cell cycle regulation, ensuring that DNA replication and repair are tightly coordinated with cell cycle progression.

Regulation

The activity and function of RPA are regulated by post-translational modifications (PTMs), such as phosphorylation. Phosphorylation of RPA2, in particular, is a critical regulatory mechanism during the cell cycle and in response to DNA damage. These modifications can alter RPA's affinity for DNA and its interactions with other proteins, modulating its role in DNA metabolism.

Clinical Significance

Given its central role in DNA replication and repair, RPA is a key player in maintaining genomic stability. Dysregulation of RPA function or expression is linked to cancer development and progression, as it can lead to increased mutation rates and genomic instability. Furthermore, RPA is a target for the development of novel cancer therapeutics, with strategies aimed at inhibiting its function to sensitize cancer cells to DNA-damaging agents.

Research Tools

RPA is also an important tool in molecular biology research. It is used in various assays to study DNA-protein interactions, DNA repair pathways, and the mechanisms of replication and recombination.