The BARD1 C-terminal domain structure and interactions with polyadenylation factor CstF-50

Area of Science:

• Biochemistry
• Molecular Biology
• Structural Biology

Background:

• The BARD1 protein interacts with BRCA1 and CstF-50.
• BARD1 plays a role in mRNA processing and RNA polymerase II stability following DNA damage.
• Understanding BARD1's structure is crucial for its function in DNA damage response.

Purpose of the Study:

• To characterize the structural biochemistry of BARD1 responsible for CstF-50 binding.
• To elucidate the role of BARD1's ankyrin (ANK) and BRCT domains in CstF-50 interaction.
• To investigate the structural flexibility of BARD1 and its implications in DNA damage signaling.

Main Methods:

• X-ray crystallography to determine the structure of the BARD1 BRCT domain.
• Small-angle X-ray scattering (SAXS) and limited proteolysis to assess domain flexibility.
• Protein pull-down assays with BARD1 deletion mutants to map CstF-50 binding sites.

Main Results:

• The BARD1 BRCT domain possesses a unique binding pocket unsuitable for phosphopeptide interactions.
• The ANK and BRCT domains are connected by a flexible linker, allowing variable orientations.
• CstF-50 binds to the ANK-BRCT linker region of BARD1, independent of the ANK or BRCT domains themselves.
• BARD1's structural plasticity facilitates the formation of diverse protein complexes for DNA damage signaling.

Conclusions:

• BARD1's flexible ANK-BRCT linker is key to its ability to integrate DNA damage signals.
• The BARD1 C-terminus acts as a adaptable hub, connecting DNA damage response pathways to RNA polymerase.
• BARD1's unique structure enables its multifaceted roles in apoptosis, p53 stabilization, and mRNA processing.