Determination of in vivo RNA structure in low-abundance transcripts

Chun Kit Kwok; Yiliang Ding; Yin Tang; Sarah M Assmann; Philip C Bevilacqua

doi:10.1038/ncomms3971

Determination of in vivo RNA structure in low-abundance transcripts

Nat Commun. 2013:4:2971. doi: 10.1038/ncomms3971.

Authors

Chun Kit Kwok¹, Yiliang Ding², Yin Tang³, Sarah M Assmann⁴, Philip C Bevilacqua⁵

Affiliations

¹ 1] Department of Chemistry, Pennsylvania State University, University Park, Pennsylvania 16802, USA [2] Center for RNA Molecular Biology, Pennsylvania State University, University Park, Pennsylvania 16802, USA [3].
² 1] Department of Chemistry, Pennsylvania State University, University Park, Pennsylvania 16802, USA [2] Center for RNA Molecular Biology, Pennsylvania State University, University Park, Pennsylvania 16802, USA [3] Department of Biology, Pennsylvania State University, University Park, Pennsylvania 16802, USA [4].
³ 1] Center for RNA Molecular Biology, Pennsylvania State University, University Park, Pennsylvania 16802, USA [2] Department of Biology, Pennsylvania State University, University Park, Pennsylvania 16802, USA [3] Bioinformatics and Genomics Graduate Program, Pennsylvania State University, University Park, Pennsylvania 16802, USA.
⁴ 1] Center for RNA Molecular Biology, Pennsylvania State University, University Park, Pennsylvania 16802, USA [2] Department of Biology, Pennsylvania State University, University Park, Pennsylvania 16802, USA [3] Bioinformatics and Genomics Graduate Program, Pennsylvania State University, University Park, Pennsylvania 16802, USA [4] Plant Biology Graduate Program, Pennsylvania State University, University Park, Pennsylvania 16802, USA.
⁵ 1] Department of Chemistry, Pennsylvania State University, University Park, Pennsylvania 16802, USA [2] Center for RNA Molecular Biology, Pennsylvania State University, University Park, Pennsylvania 16802, USA [3] Plant Biology Graduate Program, Pennsylvania State University, University Park, Pennsylvania 16802, USA.

PMID: 24336128
DOI: 10.1038/ncomms3971

Abstract

RNA structure plays important roles in diverse biological processes. However, the structures of all but the few most abundant RNAs are presently unknown in vivo. Here we introduce DMS/SHAPE-LMPCR to query the in vivo structures of low-abundance transcripts. DMS/SHAPE-LMPCR achieves attomole sensitivity, a 100,000-fold improvement over conventional methods. We probe the structure of low-abundance U12 small nuclear RNA (snRNA) in Arabidopsis thaliana and provide in vivo evidence supporting our derived phylogenetic structure. Interestingly, in contrast to mammalian U12 snRNAs, the loop of the SLIIb in U12 snRNA is variable among plant species, and DMS/SHAPE-LMPCR determines it to be unstructured. We reveal the effects of proteins on 25S rRNA, 5.8S rRNA and U12 snRNA structure, illustrating the critical importance of mapping RNA structure in vivo. Our universally applicable method opens the door to identifying and exploring the specific structure-function relationships of the multitude of low-abundance RNAs that prevail in living cells.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Arabidopsis / genetics*
Base Sequence
Chloroplasts / genetics
Evolution, Molecular
Gene Expression Profiling
Genetic Techniques
Humans
Introns
Molecular Sequence Data
Nucleic Acid Conformation*
Phylogeny
RNA Splicing
RNA, Messenger / chemistry
RNA, Ribosomal / chemistry
RNA, Ribosomal, 5.8S / chemistry
RNA, Small Nuclear / chemistry*
Sequence Homology, Nucleic Acid
Species Specificity

Substances

RNA, Messenger
RNA, Ribosomal
RNA, Ribosomal, 5.8S
RNA, Small Nuclear
U12 small nuclear RNA
RNA, ribosomal, 25S