The Shine-Dalgarno sequence of riboswitch-regulated single mRNAs shows ligand-dependent accessibility bursts.

Abstract:

:In response to intracellular signals in Gram--negative bacteria, translational riboswitches--commonly embedded in messenger RNAs (mRNAs)-regulate gene expression through inhibition of translation initiation. It is generally thought that this regulation originates from occlusion of the Shine-Dalgarno (SD) sequence upon ligand binding; however, little direct evidence exists. Here we develop Single Molecule Kinetic Analysis of RNA Transient Structure (SiM-KARTS) to investigate the ligand-dependent accessibility of the SD sequence of an mRNA hosting the 7-aminomethyl-7-deazaguanine (preQ1)-sensing riboswitch. Spike train analysis reveals that individual mRNA molecules alternate between two conformational states, distinguished by 'bursts' of probe binding associated with increased SD sequence accessibility. Addition of preQ1 decreases the lifetime of the SD's high-accessibility (bursting) state and prolongs the time between bursts. In addition, ligand-jump experiments reveal imperfect riboswitching of single mRNA molecules. Such complex ligand sensing by individual mRNA molecules rationalizes the nuanced ligand response observed during bulk mRNA translation.

journal_name

Nat Commun

journal_title

Nature communications

authors

Rinaldi AJ,Lund PE,Blanco MR,Walter NG

doi

10.1038/ncomms9976

subject

Has Abstract

pub_date

2016-01-19 00:00:00

pages

8976

issn

2041-1723

pii

ncomms9976

journal_volume

7

pub_type

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