Consistent global structures of complex RNA states through multidimensional chemical mapping

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Abstract

Accelerating discoveries of noncoding RNA (ncRNA) in myriad biological processes pose major challenges to structural and functional analysis. Despite progress in secondary structure modeling, high-throughput methods have generally failed to determine ncRNA tertiary structures, even at the 1-nm resolution that enables visualization of how helices and functional motifs are positioned in three dimensions. We report that integrating a new method called MOHCA-seq (Multiplexed •OH Cleavage Analysis with paired-end sequencing) with mutate-and-map secondary structure inference guides Rosetta 3D modeling to consistent 1-nm accuracy for intricately folded ncRNAs with lengths up to 188 nucleotides, including a blind RNA-puzzle challenge, the lariat-capping ribozyme. This multidimensional chemical mapping (MCM) pipeline resolves unexpected tertiary proximities for cyclic-di-GMP, glycine, and adenosylcobalamin riboswitch aptamers without their ligands and a loose structure for the recently discovered human HoxA9D IRES regulon. MCM offers a sequencing-based route to uncovering ncRNA 3D structure, applicable to functionally important but potentially heterogeneous states.

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Clarence Yu Cheng

Department of Biochemistry, Stanford University, Stanford, United States

Competing interests
The authors declare that no competing interests exist.
Fang-Chieh Chou

Department of Biochemistry, Stanford University, Stanford, United States

Competing interests
The authors declare that no competing interests exist.
Wipapat Kladwang

Department of Biochemistry, Stanford University, Stanford, United States

Competing interests
The authors declare that no competing interests exist.
Siqi Tian

Department of Biochemistry, Stanford University, Stanford, United States

Competing interests
The authors declare that no competing interests exist.
Pablo Cordero

Biomedical Informatics Program, Stanford University, Stanford, United States

Competing interests
The authors declare that no competing interests exist.
Rhiju Das

Department of Biochemistry, Stanford University, Stanford, United States

For correspondence
rhiju@stanford.edu

Competing interests
The authors declare that no competing interests exist.

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This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.