Dual-utility NLS drives RNF169-dependent DNA damage responses

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• DNA double-strand breaks (DSBs) trigger cell cycle checkpoints via proteins like 53BP1 and RAP80.
• High-fidelity DNA repair is essential for genomic stability but can be hindered by mediator protein deposition.
• Ring finger protein 169 (RNF169) regulates DNA damage response by limiting mediator protein accumulation.

Purpose of the Study:

• To elucidate the mechanism by which RNF169 functions in the nucleus.
• To investigate the role of RNF169's nuclear localization signal (NLS) and its interaction with USP7.
• To determine the impact of USP7-RNF169 complex disruption on DNA repair and cellular sensitivity.

Main Methods:

• Utilized structural biology (crystal structure) to guide experiments.
• Designed experiments to uncouple USP7 binding from RNF169 nuclear import.
• Assessed RNF169 stability, DSB repair fidelity, and cellular sensitivity to PARP inhibition.

Main Results:

• RNF169's NLS mediates nuclear import and stability through direct interaction with USP7.
• Perturbing the USP7-RNF169 complex destabilized RNF169 and compromised high-fidelity DSB repair.
• Disruption of the complex led to hypersensitivity to poly (ADP-ribose) polymerase (PARP) inhibition.
• USP7 and RNF169 expression levels were positively correlated in breast cancer specimens.

Conclusions:

• RNF169 employs an NLS-mediated bipartite mechanism for nuclear function in DNA damage response.
• The USP7-RNF169 interaction is critical for maintaining RNF169 stability and promoting accurate DNA repair.
• This interaction represents a potential therapeutic target for enhancing cancer treatment efficacy.