Cryo-EM structures reveal transcription initiation steps by yeast mitochondrial RNA polymerase

Brent De Wijngaert; Shemaila Sultana; Chhaya Dharia; Hans Vanbuel; Jiayu Shen; Daniel vasilchuk; Sergio E Martinez; Eaazhisai Kandiah; Smita S Patel; Kalyan Das

Comparison of PIC and IC shows little shift of upstream promoter DNA and interacting structural elements (including ICH, specificity loop, and thumb of RPO41 and NT-groove of MTF1) during PIC to IC transition. In contrast, the downstream DNA and interacting Cterminal domain of RPO41 undergo large conformational changes including fingers closing (Movie S1). During the transition, the upstream DNA from position -1 onward is locked in the MTF1 NT-groove while the template strand undergoes a large conformational . CC-BY-NC-ND 4.0 International license author/funder. It is made available under a The copyright holder for this preprint (which was not peer-reviewed) is the . https://doi.org/10.1101/2020.04.13.038620 doi: bioRxiv preprint switching to align with the 2-mer RNA and the incoming UTPaS at the active site ( Fig. 2C; Movie S2). These unsynchronized events at two ends of the transcription bubble scrunches the NT strand into an NT-loop (Movie S3). DNA scrunching has been proposed in multi-and single-subunit RNAPs including y-mtRNAP (11-14). Our y-mtIC structure is first to capture the scrunched conformation (Fig. 2C) . The scrunched NT-loop is stabilized by ICH (H641, N642), thumb (R780 and K787), and MTF1 C-tail (M334-Y335). The looping of NT strand alters the downstream DNA track and bends it from ~60° to ~120° with respect to the upstream DNA; thus, transforming a V-shaped DNA in PIC to an U-shape in IC (Fig. S5 ).