Group I permuted intron-exon (PIE) sequences self-splice to produce circular exons

Puttaraju, M.; Been, Michael D.

doi:10.1093/nar/20.20.5357

Cited by 128 publications

(108 citation statements)

References 29 publications

(22 reference statements)

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“…Theses investigations allowed us to permute the order of the intron and exon sequence without distorting the tertiary structure of the permuted intron architecture. Puttaraju & Been (1992) first reported that circular permutation of the group I intron from both the Anabena pretRNA intron and the Tetrahymena intron generated a circular RNA exon in vitro. Another PIE from the T4 phage group I intron was later shown to be applicable for generating the circular exon (Ford & Ares, 1994).…”

Section: Group I Intron Self-splicing and The Permuted Intron-exon (Pmentioning

confidence: 99%

“…As the exon sequence does not participate in the selfsplicing reaction, the exon sequence in the PIE sequence is replaced with another foreign sequence. Based on this concept, several circular RNAs have been developed by the PIE method, i.e., the tat-activated response (TAR) RNA (Puttaraju & Been, 1995;Bohjanen et al, 1996;Bohjanen et al, 1997), rev responsive element (Puttaraju & Been, 1995), HDV ribozyme (Puttaraju et al, 1993;, tRNA (Puttaraju & Been, 1992), Bacillus subtilis PRNA , mRNA encoding GFP (Perriman & Ares, 1998), yeast actin exon (Ford & Ares, 1994), hammerhead ribozyme (Ochi et al, 2009), and streptavidin RNA aptamer (Umekage & Kikuchi, 2006, 2009a, 2009b (Table 1). Fig.…”

Section: Group I Intron Self-splicing and The Permuted Intron-exon (Pmentioning

confidence: 99%

See 1 more Smart Citation

In Vivo Circular RNA Expression by the Permuted Intron-Exon Method

Umekage¹,

Uehara²,

Fujita³

et al. 2012

Innovations in Biotechnology

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Section: Group I Intron Self-splicing and The Permuted Intron-exon (Pmentioning

confidence: 99%

Section: Group I Intron Self-splicing and The Permuted Intron-exon (Pmentioning

confidence: 99%

In Vivo Circular RNA Expression by the Permuted Intron-Exon Method

Umekage¹,

Uehara²,

Fujita³

et al. 2012

Innovations in Biotechnology

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“…Finally, either Rne protein alone or purified degradosomes preferentially cleave monophosphorylated substrates 20 -30-fold more rapidly than their triphosphorylated counterparts (4). Together, these results imply that the vectorial nature of mRNA decay is a reflection of the inherent preference of Rne for 5Ј-monophosphorylated substrates.To address whether Rne or degradosomes display 5Ј-end dependence in vivo where a circular RNA would be exposed to all the components of the degradative machinery, I have constructed chimeric RNAs in which a portion of the rpsT mRNA is embedded in a permuted group I intron, a construction that should permit its precise, autocatalytic circularization (14,15). In this work I show that circular rpsT mRNAs form efficiently and are 4 -6-fold more stable than their linear counterparts.…”

mentioning

confidence: 99%

“…To address whether Rne or degradosomes display 5Ј-end dependence in vivo where a circular RNA would be exposed to all the components of the degradative machinery, I have constructed chimeric RNAs in which a portion of the rpsT mRNA is embedded in a permuted group I intron, a construction that should permit its precise, autocatalytic circularization (14,15). In this work I show that circular rpsT mRNAs form efficiently and are 4 -6-fold more stable than their linear counterparts.…”

mentioning

confidence: 99%

Stabilization of Circular rpsT mRNA Demonstrates the 5′-End Dependence of RNase E Action in Vivo

Mackie

2000

Journal of Biological Chemistry

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RNase E is the major intracellular endonuclease in Escherichia coli. Its ability to cleave susceptible substrates in vitro depends on both the cleavage site itself and the availability of an unstructured 5 terminus. To test whether RNase E activity is 5-end-dependent in vivo in the presence of all the components of the RNA degradative machinery, a known substrate, the rpsT mRNA, has been embedded in a permuted group I intron to permit its efficient, precise circularization in E. coli. Circular rpsT mRNAs are 4 -6-fold more stable in vivo than their linear counterparts. Even partial inactivation of RNase E activity further enhances this stability 6-fold. However, the stabilization of circular rpsT mRNAs depends strongly on their efficient translation. These results show unambiguously the importance of an accessible 5-end in controlling mRNA stability in vivo and support a two-step ("looping") model for RNase E action in which the first step is end recognition and the second is actual cleavage.The degradation of mRNAs is an important, if incompletely understood, aspect of the regulation of gene expression. In Escherichia coli, the initiating step in the decay process is usually mediated by RNase E (1-3), a 5Ј-end-dependent endoribonuclease (4). In vitro, RNase E activity is conferred by a multi-enzyme complex, the degradosome (5, 6). In small RNAs, such as the ColE1 replication regulator RNA 1, and the rpsO or rpsT mRNAs encoding ribosomal proteins S15 and S20, respectively, a single RNase E cleavage is capable of inactivating the mRNA and rendering it susceptible to complete destruction to mononucleotides (reviewed in Ref. 3). In larger mRNAs this initiating endonucleolytic event can trigger a 5Ј 3 3Ј "wave" of subsequent endonucleolytic cleavages, which rapidly inactivate the entire mRNA (7). The 3Ј termini generated by RNase E cleavages are scavenged by 3Ј-5Ј-exoribonucleases (1, 3).Two features in the 5Ј-extremity of an mRNA, secondary structure and the triphosphate terminus, can control the susceptibility of the entire mRNA toward RNase E. The 5Ј-terminal stem-loop structure of the ompA mRNA is largely responsible for the atypical stability of this mRNA and can confer stability to heterologous mRNAs to which it is grafted (8, 9). Stabilization is abolished by as few as three single-stranded residues at the extreme 5Ј-end of an RNA (10). These effects of 5Ј-terminal secondary structure on mRNA stability are mediated directly though Rne (4, 11). Evidence for the critical role of the free 5Ј-end of RNA and its phosphorylation state in mRNA turnover arises from several experiments. Most notably, circular derivatives of the well characterized RNase E substrates, rpsT mRNA or 9 S RNA, are highly resistant to cleavage in vitro by Rne or degradosomes (4). An RNase E cleavage at a site 5 residues from the 5Ј-end of RNA 1 destabilizes the 103-residue 3Ј-cleavage product in vivo. However, an artificial RNA identical to the initial cleavage product but containing a triphosphorylated terminus is significantly more stable ...

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“…A circularly permuted group I intron precursor RNA consisting of end-to-end fused exons that interrupt half intron sequences, self-splices to generate a circular RNA. 81 As it was demonstrated later, foreign sequences can be integrated into the exon of such a permuted self-splicing system, thus allowing the synthesis of circRNAs with desired sequence elements. 82 Based on the permuted intron exon (PIE) method, a variety of circular RNAs up to the length of 1130 nts have been generated in vitro as well as in vivo, mostly in E.coli cells.…”

Section: Ribozymes For Rna Circularizationmentioning

confidence: 97%

In vitro circularization of RNA

Müller

Appel

2016

RNA Biology

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Over the past 2 decades, different types of circular RNAs have been discovered in all kingdoms of life, and apparently, those circular species are more abundant than previously thought. Apart from circRNAs in viroids and viruses, circular transcripts have been discovered in rodents more than 20 y ago and recently have been reported to be abundant in many organisms including humans. Their exact function remains still unknown, although one may expect extensive functional studies to follow the currently dominant research into identification and discovery of circRNA by sophisticated sequencing techniques and bioinformatics. Functional studies require models and as such methods for preparation of circRNA in vitro. Here, we will review current protocols for RNA circularization and discuss future prospects in the field.Abbreviations: AMP, adenosine monophosphate; AppRNA, 5 0 -adenylated RNA; ATP, adenosine triphosphate; bp, base pair; CEV-A, citrus exocortis viroid strain A; circRNA, circular RNA; ciRNA, circular intronic RNA; CuAAC, cupper catalyzed azide alkyne cycloaddition; EDC, ethyl-3-(3 0 -dimethylaminopropyl)-carbodiimide; HIV, human immunodeficiency virus; IEciRNA, intron-exon circular RNA; nt, nucleotide; PIE, permuted intron exon; T4 Dnl, T4 DNA Ligase; T4 Rnl, T4 RNA Ligase

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Group I permuted intron-exon (PIE) sequences self-splice to produce circular exons

Cited by 128 publications

References 29 publications

In Vivo Circular RNA Expression by the Permuted Intron-Exon Method

In Vivo Circular RNA Expression by the Permuted Intron-Exon Method

Stabilization of Circular rpsT mRNA Demonstrates the 5′-End Dependence of RNase E Action in Vivo

In vitro circularization of RNA

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