Structural dynamics of RNA polymerase II throughout the nucleotide addition cycle

Area of Science:

• Molecular Biology
• Structural Biology
• Biochemistry

Background:

• Gene expression relies on RNA polymerase II (RNAPII) nucleotide addition cycles (NACs).
• Understanding the complete mechanism of NAC has been hindered by the absence of pre- and post-catalysis intermediates.
• Previous studies lacked detailed structural insights into the dynamic transitions within the RNAPII elongation complex (EC).

Purpose of the Study:

• To elucidate the complete mechanistic understanding of the RNAPII NAC.
• To capture previously intractable transition intermediates of the RNAPII EC during NAC.
• To establish a comprehensive structural and dynamic framework for RNAPII NAC.

Main Methods:

• Utilized 31 Cryo-electron Microscopy structures (43 maps) of Saccharomyces cerevisiae RNAPII EC.
• Captured distinct stages of the NAC, including transition intermediates.
• Analyzed structural dynamics across the entire NAC process.

Main Results:

• Established a continuous spectrum of RNAPII EC structural dynamics during NAC, divided into substrate-induced tightening and post-catalysis relaxation phases.
• Identified allosteric conformational changes, including Trigger Loop folding and clamp closure, upon substrate binding.
• Captured the short-lived post-catalysis product state and intermediates facilitating rapid translocation.

Conclusions:

• Defined a comprehensive structural and dynamic framework for RNAPII NAC, akin to a "molecular movie".
• Revealed a fundamental principle of balancing speed and fidelity through coordinated conformational dynamics in RNAPII.
• Provided unprecedented insight into the enzyme's mechanism during nucleotide addition.