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In vivo visualization of RNA in plants cells using the λN(22) system and a GATEWAY-compatible vector series for candidate RNAs.
Plant J. 2012 Jan 23
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19 Jun 2012
Technical Advance
DOI: 10.3410/f.716748403.792202903
Methods to visualize the subcellular location of RNAs have not been used widely in plants. These authors demonstrate that the λN(22) system is effective and compare it to the previously reported MS2 system (which has been used in plants). They find that the λN(22) system has advantages over the MS2 system, and additionally they show that both can be used together, in order to label two different RNAs in one cell. The authors end their paper "It will be interesting to see if these methods can be used to identify polar-localized mRNAs in plants" - yes, indeed.
Disclosures
None declared
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McCormick S: F1000Prime Recommendation of [Schönberger J et al., Plant J 2012]. In F1000Prime, 19 Jun 2012; DOI: 10.3410/f.716748403.792202903. F1000Prime.com/716748403#eval792202903
F1000Prime Recommendations, Dissents and Comments for [Schönberger J et al., Plant J 2012]. In F1000Prime, 20 Jun 2013; F1000Prime.com/716748403
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The past decade has seen a tremendous increase in RNA research, which has demonstrated that RNAs are involved in many more processes than were previously thought. The dynamics of RNA synthesis towards their regulated activity requires the interplay of RNAs with numerous RNA binding proteins (RBPs). The localization of RNA, a mechanism for controlling translation in a spatial and temporal fashion, requires processing and assembly of RNA into transport granules in the nucleus, transport towards cytoplasmic destinations and regulation of its activity. Compared with animal model systems little is known about RNA dynamics and motility in plants. Commonly used methods to study RNA transport and localization are time-consuming, and require expensive equipment and a high level of experimental skill. Here, we introduce the λN(22) RNA stem-loop binding system for the in vivo visualization of RNA in plant cells. The λN(22) system consists of two components: the λN(22) RNA binding peptide and the corresponding box-B stem loops. We generated fusions of λN(22) to different fluorophores and a GATEWAY vector series for the simple fusion of any target RNA 5' or 3' to box-B stem loops. We show that the λN(22) system can be used to detect RNAs in transient expression assays, and that it offers advantages compared with the previously described MS2 system. Furthermore, the λN(22) system can be used in combination with the MS2 system to visualize different RNAs simultaneously in the same cell. The toolbox of vectors generated for both systems is easy to use and promises significant progress in our understanding of RNA transport and localization in plant cells.
© 2012 The Authors. The Plant Journal © 2012 Blackwell Publishing Ltd.
DOI: 10.1111/j.1365-313X.2012.04923.x
PMID: 22268772
