DSS1 is required for the stability of BRCA2

Area of Science:

• Molecular Biology
• Cancer Research
• Genetics

Background:

• DSS1 is an evolutionarily conserved acidic protein that interacts with BRCA2.
• Investigating DSS1 function in mammalian cells is challenging due to the lack of detectable endogenous DSS1 protein via standard Western blotting.

Purpose of the Study:

• To develop a method for detecting endogenous DSS1 protein in mammalian cells.
• To elucidate the functional role of DSS1 and its interaction with BRCA2 in mammalian cells.
• To explore the potential link between DSS1 dysfunction and human cancers.

Main Methods:

• Development of a modified Western blotting protocol to detect endogenous DSS1.
• Utilizing RNA interference (RNAi) to deplete DSS1 in human cell lines.
• Analysis of BRCA2 protein levels, stability, and DNA damage sensitivity following DSS1 knockdown.

Main Results:

• A novel Western blotting protocol successfully detected endogenous DSS1.
• Essentially all BRCA2 in human cell lines is associated with DSS1.
• DSS1 depletion via RNAi led to a significant decrease in BRCA2 protein levels, primarily through increased degradation.
• A BRCA2 mutant lacking the DSS1-binding domain showed unaffected stability upon DSS1 depletion.
• DSS1 depletion resulted in hypersensitivity to DNA damage, similar to BRCA2 depletion.

Conclusions:

• The stability of BRCA2 protein in mammalian cells is dependent on the presence of DSS1.
• DSS1 dysfunction (deletion, mutation, or suppressed expression) represents a potential mechanism in sporadic and familial breast and ovarian cancers, even in the absence of BRCA1/BRCA2 mutations.