RNA loop–loop interactions as dynamic functional motifs

Brunel, Christine; Marquet, Roland; Romby, Pascale; Ehresmann, Chantal

doi:10.1016/s0300-9084(02)01401-3

Cited by 130 publications

(139 citation statements)

References 190 publications

Supporting

Mentioning

137

Contrasting

Order By: Relevance

“…The antisense and attenuator loops both have YUNR [Y-pyrimidine (C,U), R-purine (A,G), N-Y, or R] motifs, which are ubiquitous in recognition loops of natural antisense gene regulation systems (26,27), and have been used as a design element in synthetic RNA regulators (10). We therefore searched for mutations that preserved these motifs, while otherwise disrupting interac- .…”

Section: Independently Acting Attenuator Variants Can Be Engineered Tmentioning

confidence: 99%

Versatile RNA-sensing transcriptional regulators for engineering genetic networks

Lucks

Mutalik

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

234

358

View full text Add to dashboard Cite

The widespread natural ability of RNA to sense small molecules and regulate genes has become an important tool for synthetic biology in applications as diverse as environmental sensing and metabolic engineering. Previous work in RNA synthetic biology has engineered RNA mechanisms that independently regulate multiple targets and integrate regulatory signals. However, intracellular regulatory networks built with these systems have required proteins to propagate regulatory signals. In this work, we remove this requirement and expand the RNA synthetic biology toolkit by engineering three unique features of the plasmid pT181 antisense-RNA-mediated transcription attenuation mechanism. First, because the antisense RNA mechanism relies on RNA-RNA interactions, we show how the specificity of the natural system can be engineered to create variants that independently regulate multiple targets in the same cell. Second, because the pT181 mechanism controls transcription, we show how independently acting variants can be configured in tandem to integrate regulatory signals and perform genetic logic. Finally, because both the input and output of the attenuator is RNA, we show how these variants can be configured to directly propagate RNA regulatory signals by constructing an RNA-meditated transcriptional cascade. The combination of these three features within a single RNA-based regulatory mechanism has the potential to simplify the design and construction of genetic networks by directly propagating signals as RNA molecules.gene networks | regulatory systems | orthogonal regulators N oncoding RNA has been found to play a central role in regulating gene expression in both prokaryotes and eukaryotes. Recently, the diverse roles of RNA-mediated regulation have become important tools for synthetic biology applications ranging from detecting metabolic state (1), balancing metabolic pathway expression (2), tightly regulating toxin genes (3), and detecting environmentally harmful chemicals (4). In particular, RNA-based genetic parts have been engineered that regulate transcription through RNA-mediated transcription factor recruitment (5, 6), transcript stability through small-molecule-mediated ribozyme cleavage (1, 7) and siRNA targeted degradation (8), and translation through cis-acting mRNA conformational changes (9) and trans-acting antisense RNA-mRNA interactions (10,11).This wide array of RNA function is beginning to be used to engineer programmable genetic circuitry required for the next level of synthetic biology applications (12). By interfacing with protein-based transcription factors and repressors, hybrid RNA∕ protein cascades have been made that perform sophisticated logic evaluation (8) and even count extracellular events (13). Much like previous work on protein-based cascades (14), protein regulators propagate the signal between different levels of the hybrid cascades. This makes the inner workings of these cascades complicated by the many interconversions between mRNA and protein that must take place. This not only incre...

show abstract

Section: Independently Acting Attenuator Variants Can Be Engineered Tmentioning

confidence: 99%

Versatile RNA-sensing transcriptional regulators for engineering genetic networks

Lucks

Mutalik

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

234

358

View full text Add to dashboard Cite

show abstract

“…21 They have, therefore, been much better characterized for RNA, both in terms of their structure [22][23][24] and mechanical properties, 25,26 than for DNA. This structural knowledge has even been exploited in structural RNA nanotechnology, where kissing loop interactions have also been used as a means to join RNA components with a well-defined geometry.…”

Section: Introductionmentioning

confidence: 99%

The effect of topology on the structure and free energy landscape of DNA kissing complexes

Romano

Hudson

Doye

et al. 2012

The Journal of Chemical Physics

View full text Add to dashboard Cite

We use a recently developed coarse-grained model for DNA to study kissing complexes formed by hybridization of complementary hairpin loops. The binding of the loops is topologically constrained because their linking number must remain constant. By studying systems with linking numbers −1, 0, or 1 we show that the average number of interstrand base pairs is larger when the topology is more favourable for the right-handed wrapping of strands around each other. The thermodynamic stability of the kissing complex also decreases when the linking number changes from −1 to 0 to 1. The structures of the kissing complexes typically involve two intermolecular helices that coaxially stack with the hairpin stems at a parallel four-way junction.

show abstract

“…A kissing interaction, a basic type of RNA tertiary contact, is the base-pairing formed by complementary sequences in the apical loops of two hairpins (5). Intramolecular kissing complexes have been found in many RNA structures, ranging from 75-nt tRNAs (6,7) to megadalton ribosomes (8); intermolecular kissing interactions also are critical for many biological processes (reviewed in ref.…”

mentioning

confidence: 99%

“…Intramolecular kissing complexes have been found in many RNA structures, ranging from 75-nt tRNAs (6,7) to megadalton ribosomes (8); intermolecular kissing interactions also are critical for many biological processes (reviewed in ref. 5), such as dimerization of retroviral genomic RNAs (9,10). The simplest kissing interaction is formed between a pair of hairpins each with a GACG tetraloop (11).…”

mentioning

confidence: 99%

Unusual mechanical stability of a minimal RNA kissing complex

Li¹,

Bustamante

Tinoco³

2006

Proc. Natl. Acad. Sci. U.S.A.

146

View full text Add to dashboard Cite

By using optical tweezers, we have investigated the mechanical unfolding of a minimal kissing complex with only two G⅐C base pairs. The loop-loop interaction is exceptionally stable; it is disrupted at forces ranging from 7 to 30 pN, as compared with 14 -20 pN for unfolding hairpins of 7 and 11 bp. By monitoring unfolding͞ folding trajectories of single molecules, we resolved the intermediates, measured their rate constants, and pinpointed the ratelimiting steps. The two hairpins unfold only after breaking the intramolecular kissing interaction, and the kissing interaction forms only after the folding of the hairpins. At forces that favor the unfolding of the hairpins, the entire RNA structure is kinetically stabilized by the kissing interaction, and extra work is required to unfold the metastable hairpins. The strong mechanical stability of even a minimal kissing complex indicates the importance of such loop-loop interactions in initiating and stabilizing RNA dimers in retroviruses.mechanical unfolding ͉ optical tweezers ͉ RNA dimerization ͉ single molecule ͉ RNA folding T ertiary interactions enable RNA to form long-range contacts and thereby form complex structures. However, thermodynamic and kinetic information for these interactions is scarce (1). Recent developments in single-molecule techniques allow a close look at the folding of individual RNA molecules (2-4). Particularly, the application of force to single-ribozyme molecules by optical tweezers (3) revealed the detailed unfolding pathways of this 390-nt RNA in nondenaturing solutions at physiological temperatures. It remains a challenge, however, to study the kinetics of individual steps in folding an RNA with complicated tertiary structures.A kissing interaction, a basic type of RNA tertiary contact, is the base-pairing formed by complementary sequences in the apical loops of two hairpins (5). Intramolecular kissing complexes have been found in many RNA structures, ranging from 75-nt tRNAs (6, 7) to megadalton ribosomes (8); intermolecular kissing interactions also are critical for many biological processes (reviewed in ref. 5), such as dimerization of retroviral genomic RNAs (9, 10). The simplest kissing interaction is formed between a pair of hairpins each with a GACG tetraloop (11). The third and fourth nucleotides in the loop form two G⅐C base pairs with their counterparts from the other hairpin. This minimal kissing interaction was initially found in the genomic RNA of Moloney murine leukemia virus (MMLV), an extensively studied retrovirus (12) and one of the most used vectors for gene therapy (13). The 5Ј UTR of the MMLV genomic RNA contains four closely spaced stem loops (SL-A-SL-D) (14), each of which is capable of forming a kissing interaction with its counterpart in another copy of the genomic RNA (11,15,16). This region, including the two hairpins with GACG loops (SL-C and SL-D), serves both as the RNA dimerization initiation site (DIS) and as the RNA encapsidation signal () (17, 18).The kissing hairpins are evolutionarily conserved in the...

show abstract

RNA loop–loop interactions as dynamic functional motifs

Cited by 130 publications

References 190 publications

Versatile RNA-sensing transcriptional regulators for engineering genetic networks

Versatile RNA-sensing transcriptional regulators for engineering genetic networks

The effect of topology on the structure and free energy landscape of DNA kissing complexes

Unusual mechanical stability of a minimal RNA kissing complex

Contact Info

Product

Resources

About