Multi-domain conformational selection underlies pre-mRNA splicing regulation by U2AF.

Area of Science:

• Molecular Biology
• Biochemistry
• Genetics

Background:

• Multi-domain proteins are crucial for cellular functions, recognizing specific ligands through their independent modules.
• Understanding how multiple domains interact dynamically during ligand recognition is essential.
• The U2 auxiliary factor 65 (U2AF65) protein plays a key role in pre-mRNA splicing by recognizing the polypyrimidine tract RNA.

Purpose of the Study:

• To elucidate the molecular mechanisms by which U2AF65 recognizes the 3'-splice-site-associated polypyrimidine tract RNA.
• To investigate the role of U2AF65's domain dynamics in pre-mRNA splicing regulation.

Main Methods:

• Studied the molecular mechanisms of polypyrimidine tract RNA recognition by U2AF65.
• Utilized structural and functional assays to characterize U2AF65 conformations and RNA binding.
• Investigated the impact of mutations on conformational equilibrium and splicing activity.

Main Results:

• U2AF65's tandem RNA recognition motif domains adopt distinct closed and open conformations based on polypyrimidine tract affinity.
• RNA sequence variations induce population shifts between these conformations.
• The conformational equilibrium acts as a molecular rheostat, correlating RNA features to U2 snRNP recruitment efficiency.

Conclusions:

• U2AF65 employs a conformational selection mechanism for degenerate RNA motif recognition.
• This mechanism quantitatively links polypyrimidine tract characteristics to spliceosome assembly efficiency.
• Cooperative multi-domain conformational selection may be a general principle in molecular recognition.