Ubx5-Cdc48 assists the protease Wss1 at DNA-protein crosslink sites in yeast

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• DNA-protein crosslinks (DPCs) are DNA lesions that compromise genome stability.
• Yeast proteases like Wss1, 26S proteasome, and Ddi1 maintain genome integrity by processing DNA-bound proteins.
• The AAA ATPase Cdc48/p97 is implicated in DNA-protein complex clearance, but its role in DPC proteolysis is not fully understood.

Purpose of the Study:

• To investigate the role of the Cdc48 adaptor Ubx5 in DNA-protein crosslink (DPC) processing and genome stability.
• To elucidate the interplay between Ubx5, Cdc48, and the Wss1 protease in DPC repair.
• To determine the contribution of Ubx5-Cdc48 to the degradation of DNA-bound proteins, such as RNA polymerase II.

Main Methods:

• Utilized yeast mutants defective in DPC processing.
• Employed an inducible site-specific crosslinking system to generate and study DPCs.
• Assessed the accumulation of Ubx5 at DPC lesions in Wss1-deficient cells.
• Investigated the effect of abolishing Cdc48 binding or Ubx5 loss on DPC sensitivity.
• Examined the degradation of RNA polymerase II (RNAPII) in response to genotoxic agents.

Main Results:

• Ubx5 is detrimental in yeast mutants with impaired DPC processing.
• Ubx5 accumulates at persistent DPC lesions in the absence of Wss1, hindering DPC removal.
• Loss of Ubx5 or impaired Cdc48 binding suppresses the sensitivity of wss1∆ cells to DPC-inducing agents by promoting alternative repair pathways.
• Demonstrated cooperation between Ubx5-Cdc48 and Wss1 in the genotoxin-induced degradation of RNA polymerase II (RNAPII).

Conclusions:

• Ubx5 plays a critical role in the clearance and repair of DNA-protein crosslinks (DPCs).
• The Ubx5-Cdc48 complex collaborates with Wss1 protease to facilitate the proteolysis of specific DNA-bound proteins.
• These findings reveal a novel mechanism involving Ubx5 in maintaining genome integrity against DPC-induced damage.