Piccirilli, J. A. | Search bioRxiv

bioRxiv2021

The cardiac lncRNA Chantico directly regulates Cxcl1 chemokine transcription

Withdrawal StatementThe corresponding author has withdrawn this preprint owing to inability to reproduce some of the data, instances of inappropriate data exclusion, and loss of much of the primary experimental records/data. Specifically, Figures 2B,E,H; 3; 4; 5A,B,D; 6; S2A,C; S4A; S5; and S6A,B and attendant text contain analyses for which the primary record and/or raw data no longer exist; the analyses, where still available, suffer from inappropriate data exclusion and thus should not be construed to be an accurate reflection of the experiments. Attempts by others in the lab to repeat several of the experiments in these indicated panels have failed reproduce the presented effects, despite showing much greater precision. Therefore the authors do not wish this work to be cited as reference for the project. If you have any questions, please contact the corresponding author.

molecular biology↗

bioRxiv2021

Sub-3 Angstrom cryo-EM structure of RNA enabled by engineered homomeric self-assembly

Many functional RNAs fold into intricate and precise 3D architectures, and high-resolution structures are required to understand their underlying mechanistic principles. However, RNA structural determination is difficult. Herein, we present a nanoarchitectural strategy to enable the efficient single-particle cryogenic electron microscopy (cryo-EM) analysis of RNA-only structures. This strategy, termed RNA oligomerization-enabled cryo-EM via installing kissing-loops (ROCK), involves the engineering of target RNAs by installing kissing-loop sequences onto functionally nonessential stems for the assembly into closed homomeric nanoarchitectures. Assembly with geometric restraints leads to (1) molecular weight multiplication and (2) structural flexibility mitigation, both beneficial for cryo-EM analysis. Together with construct optimization and symmetry-expansion reconstruction, ROCK yields the cryo-EM reconstruction of the Tetrahymena group I intron at an overall resolution of 2.98 [A] (2.85 [A] resolution for the core domains), enabling the de novo model building of the complete intron RNA including previously unknown peripheral domains. When applied to smaller RNAs, ROCK readily produces modest-resolution maps, revealing the conformational rearrangement of the Azoarcus group I intron and the bound ligand in the FMN riboswitch. Our work unleashes the largely unexplored potential of cryo-EM in RNA structural studies.

biophysics↗