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The effect of replication protein A inhibition and post-translational modification on ATR kinase signaling

Matthew R Jordan1, Greg G Oakley2, Lindsey D Mayo3

  • 1Department of Medicine, Indiana University School of Medicine, Indianapolis, IN, 64202, USA.

Scientific Reports
|August 26, 2024
PubMed

Insights

New chemical RPA inhibitors block ATR signaling by preventing RPA interactions, offering a targeted approach for cancer therapeutics. These inhibitors show selectivity advantages over existing treatments.

Area of Science:

  • Molecular Biology
  • Cancer Biology
  • Biochemistry

Background:

  • The ATR kinase pathway is crucial for maintaining genome integrity by responding to DNA damage.
  • Cancer cells often exhibit intrinsic replication stress, making the ATR pathway a promising therapeutic target.
  • Replication Protein A (RPA) is essential for sensing single-stranded DNA (ssDNA) and activating ATR signaling.

Purpose of the Study:

  • To develop and characterize novel chemical inhibitors of RPA (RPAi).
  • To elucidate the mechanism of action of RPA inhibitors in blocking ATR kinase activation.
  • To investigate the impact of RPA post-translational modifications (PTMs) on ATR signaling and RPAi sensitivity.

Main Methods:

  • Biochemical reconstitution of the ATR kinase signaling pathway.
  • Development of RPA inhibitors targeting RPA-ssDNA interactions (RPA-DBi) and RPA protein-protein interactions (RPA-PPIi).
  • Assessment of ATR-dependent phosphorylation of target proteins in response to RPAi.

Main Results:

  • RPA-DBi and RPA-PPIi effectively abrogate ATR-dependent phosphorylation with greater selectivity than active site ATR inhibitors.
  • RPA PTMs, such as RPA32 phosphorylation and TopBP1 phosphorylation, stimulate ATR kinase activation.
  • RPA70 acetylation does not influence ATR phosphorylation of target proteins, and RPAi sensitivity is unaffected by RPA PTMs.

Conclusions:

  • This study reveals the mechanism by which RPA inhibitors block ATR signaling.
  • RPA PTMs can modulate ATR kinase activation but do not alter sensitivity to RPA inhibitors.
  • RPA inhibitors offer a promising strategy for developing novel anti-cancer therapeutics targeting the ATR pathway.

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