Distinct Mechanisms of Transcription Initiation by RNA Polymerases I and II

Supplemental Movie 1: Molecular Animation of the Pol I Transcription Cycle
The movie visualizes the steps of the Pol I transcription cycle as shown in Figure 4. In an initial overview slide, the parts of the movie are outlined. Structures solved by X-ray crystallography are indicated by a crystal symbol, with structures originating from cryo-EM reconstructions by an EM grid symbol (compare to Table 1). In the beginning, the main structural modules are depicted in one monomer of the Pol I crystal structure (PDB 4C2M) in 3D by rotating around the y-axis for 360° starting from the front view. The Pol I–specific bridge helix unwinding, A12.2 C-terminal domain, and expander are depicted as ribbon models prior to outlining the mode of dimerization via mutual stalk–cleft interactions. Following, the cleft contraction upon monomerization is depicted, which is achieved by a rotation of core and shelf modules around an axis that runs through the active site of the polymerase. Next, binding of Rrn3 to the stalk domain is shown in a side view, depicting how phosphorylation of a serine patch on Rrn3 prevents the interaction. Rotating back to the front view, the structure of the Pol I–Rrn3 complex is presented, to which the core factor consisting of Rrn6 (violet), Rrn7 (green), and Rrn11 (yellow) can bind on its own using its polymerase-interacting regions I, II, and III. However, when CF is bound to promoter DNA (nontemplate strand in cyan, template strand in blue), it interacts with the protrusion domain of Pol I instead of with the clamp core, forming a closed complex. The closed complex is rotated around the y-axis for 270°, pausing at an A135 side view with transparent proteins, indicating the architecture of a modeled open complex with melted DNA strands. Following melting, initial transcription yields a short hybrid (RNA appears in red). After fading out shortly, the same transition from CC to OC to ITC is repeated from a front view that shows how the Pol I modules contribute to the continuing cleft contraction, ending with the structure of a productive, fully contracted EC in which the A12.2 C-terminal domain is detached. Finally, a zoom-in shows how the bridge helix is rewound in the EC compared to the dimeric crystal structure and how the downstream DNA duplex is positioned, and the movie concludes with a zoom-out to the front view with which it began. Used PDB structures are 4C2M, 5M3M, 3TJ1, 5G5L, 5N5X, 5N5Y, 5N61, and 5M3F (11, 33–38, 97). The movie was recorded using UCSF Chimera (101). Abbreviations: CC, closed complex; CF, core factor; EC, elongation complex; EM, electron microscopy; ITC, initially transcribing complex; OC, open complex; PDB, Protein Data Bank; Pol, RNA polymerase.