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Stalling of Eukaryotic Translesion DNA Polymerases at DNA-Protein Cross-Links

Anna V Yudkina1, Evgeniy S Shilkin2, Alena V Makarova2

  • 1Siberian Branch of the Russian Academy of Sciences Institute of Chemical Biology and Fundamental Medicine, 8 Lavrentieva Ave., 630090 Novosibirsk, Russia.

Genes
|February 25, 2022
PubMed

Insights

DNA-protein cross-links (DPCs) hinder DNA replication. DNA polymerases ζ (POLζ) and η (POLη) show varying efficiency in bypassing DPCs, with POLη being the most effective at navigating these bulky adducts.

Area of Science:

  • Molecular Biology
  • Biochemistry
  • Genetics

Background:

  • DNA-protein cross-links (DPCs) are bulky DNA adducts that impede DNA replication.
  • SPRTN protease processes DPCs in human cells, activated by stalled DNA polymerases.
  • A proposed mechanism involves protein surface clashes and distortion of cross-linked proteins.

Purpose of the Study:

  • To investigate the behavior of eukaryotic translesion DNA polymerases (POLζ, POLι, POLη) on model DPC-containing DNA substrates.
  • To compare the efficiency of different polymerases in synthesizing DNA across DPCs.
  • To validate the proposed mechanism of polymerase-DPC interaction.

Main Methods:

  • Utilized model DPCs in single-stranded template, template strand, or displaced strand DNA.
  • Assessed the synthesis activity of DNA polymerases ζ (POLζ), ι (POLι), and η (POLη) on these substrates.
  • Analyzed polymerase pausing and nucleotide incorporation at DPC sites.

Main Results:

  • POLι exhibited limited synthesis on DPC-containing DNA.
  • POLζ and POLη paused at polymerase and cross-linked protein footprints.
  • POLη demonstrated the highest efficiency, incorporating nucleotides past the DPC site.
  • A DPC in the displaced strand blocked all tested polymerases.

Conclusions:

  • Translesion DNA polymerases display differential abilities to process DPCs.
  • POLη is the most efficient polymerase studied for bypassing DPCs.
  • Encountered DPC behaviors align with the protein clash and distortion model.
Keywords:
DNA polymerasesDNA replicationDNA–protein cross-linktranslesion synthesis

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