Evolution of Rev7 interactions in eukaryotic TLS DNA polymerase Polζ

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• Translesion synthesis (TLS) DNA polymerase Polζ bypasses DNA damage.
• Rev7 is a HORMA protein subunit of Polζ, crucial for its recruitment to replication forks.
• Human Rev7 (hRev7) dimerizes via its HORMA interface when bound to hRev3, a mechanism conserved in HORMA proteins.

Purpose of the Study:

• To investigate the dimerization of Saccharomyces cerevisiae Rev7 (scRev7) in the context of Polζ.
• To compare the Rev7-Rev3 interaction and dimerization mechanisms between yeast and humans.

Main Methods:

• Biophysical studies (e.g., surface plasmon resonance) to measure binding affinities.
• Analysis of cryo-electron microscopy (cryo-EM) structures.
• Validation of Rev7-binding motifs (RBMs) in scRev3.

Main Results:

• Two adjacent RBMs in scRev3 bind scRev7 with significantly different affinities (two orders of magnitude).
• The scRev7:Rev3 complex exists in a 2:1 stoichiometry in solution.
• scRev7 does not form dimers in solution, either independently or when bound to scRev3.
• The scRev7 dimer observed in cryo-EM structures is likely induced by interactions with other Polζ subunits.

Conclusions:

• Rev7 homodimerization via the HORMA interface is not conserved in yeast Polζ.
• The mechanism of Rev7 dimerization appears to have evolved later in eukaryotic evolution.
• Rev7 homodimerization is not a universal requirement for Polζ function.