Zinc-finger protein CNBP alters the 3-D structure of lncRNA Braveheart in solution

Doo Nam Kim,Scott P Hennelly,Karissa Y Sanbonmatsu,Tyler Mrozowich,Trushar R Patel,Bernhard C Thiel,Ivo L Hofacker

doi:10.1038/s41467-019-13942-4

Abstract

Long non-coding RNAs (lncRNAs) constitute a significant fraction of the transcriptome, playing important roles in development and disease. However, our understanding of structure-function relationships for this emerging class of RNAs has been limited to secondary structures. Here, we report the 3-D atomistic structural study of epigenetic lncRNA, Braveheart (Bvht), and its complex with CNBP (Cellular Nucleic acid Binding Protein). Using small angle X-ray scattering (SAXS), we elucidate the ensemble of Bvht RNA conformations in solution, revealing that Bvht lncRNA has a well-defined, albeit flexible 3-D structure that is remodeled upon CNBP binding. Our study suggests that CNBP binding requires multiple domains of Bvht and the RHT/AGIL RNA motif. We show that RHT/AGIL, previously shown to interact with CNBP, contains a highly flexible loop surrounded by more ordered helices. As one of the largest RNA-only 3-D studies, the work lays the foundation for future structural studies of lncRNA-protein complexes.

Highlights

Long non-coding RNAs constitute a significant fraction of the transcriptome, playing important roles in development and disease
In the epigenetics and pharmaceutical communities, there has been great interest in the question: do Long non-coding RNAs (lncRNAs) have well-defined structures? It has not been clear whether the majority of lncRNAs are disordered, extended or compact[46]
Due to their large structures (200–100,000 nts) and dynamic binding function with partner molecules, it is sometimes assumed that lncRNA structures are generally too disordered, or highly flexible to be studied by high-resolution structural determination methods

Summary

Introduction

Long non-coding RNAs (lncRNAs) constitute a significant fraction of the transcriptome, playing important roles in development and disease. Akin to early studies of the ribosome, these secondary structures provide the framework for understanding structure–function relationships, producing valuable information like local modularity; chemical probing studies yield little information about the overall 3-dimensional (3-D) structure[11] As their name suggests, long noncoding RNAs are often large, making their preparation and purification for in vitro studies very challenging. Many biologically important RNAs have dynamically changing conformations, making structure determination challenging[2,15] It has been shown, using chemical probing, that several lncRNAs and portions of lncRNAs adopt well-organized, modular secondary structures[3,4,5,7,9].

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Results

Conclusion