Widespread epistasis shapes RNA polymerase II active site function and evolution

Area of Science:

• Molecular Biology
• Structural Biology
• Biochemistry

Background:

• Multi-subunit RNA polymerases are essential for transcription across all life forms.
• The trigger loop is a dynamic, conserved active site domain critical for transcription cycle steps.
• Mutations in the RNA polymerase II trigger loop lead to distinct phenotypes affecting catalysis.

Purpose of the Study:

• To structurally map the interaction landscape of the RNA polymerase II trigger loop.
• To understand how residue interactions influence trigger loop function and transcription.
• To investigate the impact of mutations on the RNA polymerase II active site network.

Main Methods:

• Deep mutational scanning was employed on Saccharomyces cerevisiae RNA polymerase II.
• A structural genetics approach was used to analyze residue interactions.
• The study focused on the trigger loop and its surrounding domains.

Main Results:

• The research identified key residue interactions within and around the RNA polymerase II trigger loop.
• A comprehensive "interaction landscape" of the trigger loop was determined.
• Connections between trigger loop residues and other domains were revealed.

Conclusions:

• Trigger loop function is intrinsically linked to its specific enzymatic context.
• Understanding these interactions provides insights into transcription regulation.
• This work highlights the importance of residue networks in enzyme activity.