Excision of Group II Introns as Circles

Murray, Heather L.; Mikheeva, Svetlana A.; Coljee, Vincent W.; Turczyk, Brian M.; Donahue, William F.; Bar-Shalom, Avital; Jarrell, Kevin A.

doi:10.1016/s1097-2765(01)00300-8

Cited by 57 publications

(75 citation statements)

References 29 publications

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“…It is highly significant that the RT-PCR products derived from intron forms with linked 5Ј and 3Ј ends generated by ␥ЈG mutants contained an abnormality, in that a G residue was incorporated instead of a C nucleotide when reverse transcriptase encountered the terminal G residue of the intron at the junction. These findings suggest the presence of a 2Ј-5Ј junction and may therefore represent intron circles (14). It therefore seems that these intron forms with linked ends produced by RmInt1 may result as suggested previously (14) from intron molecules that undergo cleavage at the 3Ј splice site rather than the first step of the branching pathway, by a trans-splicing reaction triggered by 5Ј exon molecules generated by the so-called spliced exon reopening reaction.…”

Section: Discussionsupporting

confidence: 57%

“…linkage (14). Thus, one possible interpretation of our data is that the extension product observed in the EBS3C mutant could be mostly the larger 98-nt product and that the mutant intron might excised in vivo as putative intron circles.…”

Section: Detection Of Rmint1 Products In Vivo Bymentioning

confidence: 71%

“…We have shown previously that primer extension assays using RNA and RNP particles from S. meliloti cells expressing the wild-type intron revealed two extension products that differ by one nucleotide (97-98 nt) relative to the position expected for the 5Ј end of the intron, with the shorter product as the major one (16). Interestingly, primer extension performed on a circular intron has been reported to result in two extension products that differ by one nucleotide, with the longer product as the major one (14). Furthermore, reverse transcriptase is known to pause after reading through the 2Ј-5Ј linkage in a nonlariat RNA (28), whereas with lariat RNA, the pause occurs before reading through the 2Ј-5Ј FIGURE 3.…”

Section: Detection Of Rmint1 Products In Vivo Bymentioning

confidence: 99%

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Relevance of the Branch Point Adenosine, Coordination Loop, and 3′ Exon Binding Site for in Vivo Excision of the Sinorhizobium meliloti Group II Intron RmInt1

Molina-Sánchez¹,

Barrientos‐Durán²,

Toro

2011

Journal of Biological Chemistry

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Excision of the bacterial group II intron RmInt1 has been demonstrated in vivo, resulting in the formation of both intron lariat and putative intron RNA circles. We show here that the bulged adenosine in domain VI of RmInt1 is required for splicing via the branching pathway, but branch site mutants produce small numbers of RNA molecules in which the first G residue of the intron is linked to the last C residue. Mutations in the coordination loop in domain I reduced splicing efficiency, but branched templates clearly predominated among splicing products. We also found that a single substitution at the EBS3 position (G329C), preventing EBS3-IBS3 pairing, resulted in the production of 50 to 100 times more RNA molecules in which the 5 and 3 extremities were joined. We provide evidence that these intron molecules may correspond to both, intron circles linked by a 2-5 phosphodiester bond, and tandem, head-to-tail intron copies.Group II introns are large catalytic RNAs with a conserved secondary structure consisting of six domains arranged around a central wheel (1). Group II introns may self-splice via several different pathways (Fig. 1). The main splicing pathway in group II introns involves two sequential trans-esterification reactions. In the first, the 2Ј-hydroxyl of a bulged adenosine near the end of the intron, in domain VI, triggers a nucleophilic attack on the 5Ј splice site. In the second reaction, the 2Ј-5Ј branched product is released as a lariat when the free 3Ј-OH at the end of the 5Ј exon attacks the 3Ј splice junction joining the two exons (2-5). In ai5␥, a yeast mitochondrial group IIB intron, the functional group II intron branch site consists of the bulged adenosine, the wobble pairs (G-U) flanking this residue and, possibly, the purine-enriched sequence in domain VI on the strand opposite the bulged nucleotide (6 -8). Recent studies have reported the presence, within domain I of group IIB introns, of a branch site receptor known as a coordination loop (9), which is required for trans-esterification (8), but these findings have been called into question (10). This coordination loop has been identified as a key element for exon ligation, as it positions the exon and intron binding sequences, the EBS3-IBS3 base pair and the EBS1-IBS1 helix, in an appropriate conformation for reaction (9). There is also an alternative splicing pathway, the first step of which involves hydrolysis. The ligated exons and linear intron are then generated in a standard second step (11-13). An additional excision reaction, in which the intron is excised as a putative circle, has also been proposed for some group II introns (14 -16). In this case, prior release of the 3Ј exon from the precursor molecules, presumably via a transsplicing mechanism, seems to be required for the generation of a putative 2Ј-5Ј ligated circular intron. The origin and function of such intron circles in vivo remains unknown.RmInt1 is a mobile subclass IIB3 intron (17). It has also been assigned to phylogenetic bacterial class D (17, 18) on the basis o...

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Section: Discussionsupporting

confidence: 57%

Section: Detection Of Rmint1 Products In Vivo Bymentioning

confidence: 71%

Section: Detection Of Rmint1 Products In Vivo Bymentioning

confidence: 99%

See 1 more Smart Citation

Relevance of the Branch Point Adenosine, Coordination Loop, and 3′ Exon Binding Site for in Vivo Excision of the Sinorhizobium meliloti Group II Intron RmInt1

Molina-Sánchez¹,

Barrientos‐Durán²,

Toro

2011

Journal of Biological Chemistry

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“…During the formation of circRNAs either 2 0 -5 0 or 3 0 -5 0 linkages have been detected. 8,9 The resulting molecular "rings" or RNA circles resist degradation by exoribonucleases that require free termini and/or may have increased melting point in comparison to linear nucleic acid molecules. Circularization of RNA molecules may thus drastically influence the structure and/or shape of these molecules, presumably reflecting structural constraints brought about by circularization.…”

Section: Introductionmentioning

confidence: 99%

High-throughput sequencing reveals circular substrates for an archaeal RNA ligase

et al. 2017

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It is only recently that the abundant presence of circular RNAs (circRNAs) in all kingdoms of Life, including the hyperthermophilic archaeon Pyrococcus abyssi, has emerged. This led us to investigate the physiologic significance of a previously observed weak intramolecular ligation activity of Pab1020 RNA ligase. Here we demonstrate that this enzyme, despite sharing significant sequence similarity with DNA ligases, is indeed an RNA-specific polynucleotide ligase efficiently acting on physiologically significant substrates. Using a combination of RNA immunoprecipitation assays and RNA-seq, our genome-wide studies revealed 133 individual circRNA loci in P. abyssi. The large majority of these loci interacted with Pab1020 in cells and circularization of selected C/D Box and 5S rRNA transcripts was confirmed biochemically. Altogether these studies revealed that Pab1020 is required for RNA circularization. Our results further suggest the functional speciation of an ancestral NTase domain and/or DNA ligase toward RNA ligase activity and prompt for further characterization of the widespread functions of circular RNAs in prokaryotes. Detailed insight into the cellular substrates of Pab1020 may facilitate the development of new biotechnological applications e.g. in ligation of preadenylated adaptors to RNA molecules.

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“…Some of the well-known circular RNAs so far were Group I and II introns ( Vicens and Cech, 2009;Murray et al, 2001), viroids and satellite RNAs (Hammann and Steger, 2012) and special cases of skipped exons (Zaphiropoulos, 1997; Burd et al, 2010;Capel et al, 1993). Due to their specialized nature they were not thought to be of general physiological relevance.…”

Section: Circular Rna and Sine Ecombination In Entamoeba Histolyticamentioning

confidence: 99%

Circular Noncoding Ribosomal Transcripts and Recombination of Retrotransposons: Two Charming Secrets Revealed by a Less-Explored Human Parasite

Bhattacharya¹

2015

Proceedings of the Indian National Science Academy

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Parasites, through their diverse cellular metabolic pathways, help us to appreciate the different directions in which evolution works, while maintaining a unified theme. To illustrate this I describe our recent observations with two basic processes, namely regulation of ribosomal RNA transcription, and the biology of retrotransposition in Entamoeba histolytica, a highly prevalent protozoan parasite that causes amebiasis.Ribosomal RNA synthesis is generally tightly regulated in response to growth rate such that cells subjected to growth stress shut down their rRNA transcription. In E. histolytica we observed that upon growth stress, rRNA synthesis did not shut down. Instead, unprocessed pre-rRNA accumulated to high levels along with a novel class of circular RNAs derived from the 5'-external transcribed spacer (etsRNA). The etsRNA can self circularize in vitro, a property not previously known in spacer RNAs. We hypothesize that circular etsRNAs would escape exonucleolytic decay and inhibit pre-rRNA processing, possibly by titrating away the processing factors which normally bind to them.In the study on retrotransposition of non-long terminal repeat retrotransposons (EhLINEs/SINEs), we successfully mobilized EhSINE in a cell-line made retrotransposition-competent by transfection with multiple constructs to express the polypeptides required for retrotransposition, a first for any protozoan parasite. While tracking retrotransposition of a marked SINE copy we found that the newly retrotransposed SINEs had undergone high-frequency recombination, presumably due to the known ability of reverse transcriptase to perform template jumping. Such recombination has not been reported for retrotransposons, and may be important in generating sequence polymorphism.Key Words: Entamoeba histolytica; Circular Non Coding RNA; Ribosomal RNA Spacer; Ehsine; Non LTR Retrotransposon; SINE Recombination IntroductionEntamoeba histolytica, a protozoan parasite, is classically defined as the causative agent of the disease Amoebiasis. It resides in the human colon along with a number of nonpathogenic species of Entamoeba. The estimates available world-wide state that 500 million persons are infected with Entamoeba spp., with about 50 million cases of invasive amoebiasis per year; resulting in about 100,000 deaths (WHO/PAHO/UNESCO report, 1997). The parasite is directly transmitted through the faecal-oral route without the need of a vector system. It therefore flourishes where humans live in crowded conditions with poor sanitation. It continues to be a major public 432 Sudha Bhattacharya health problem in India due to recurrent infections caused by the high parasite load in our environment, and lack of effective immunity in infected individuals. The morbidity caused by the intestinal form of the disease results in great economic loss, although mortality is reduced, thanks to the freely-available and over-used anti-amoebic drug, metronidazole (Upcroft and Upcroft, 2001). The parasite also invades the liver and other organs, causing abscesses ...

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Excision of Group II Introns as Circles

Cited by 57 publications

References 29 publications

Relevance of the Branch Point Adenosine, Coordination Loop, and 3′ Exon Binding Site for in Vivo Excision of the Sinorhizobium meliloti Group II Intron RmInt1

Relevance of the Branch Point Adenosine, Coordination Loop, and 3′ Exon Binding Site for in Vivo Excision of the Sinorhizobium meliloti Group II Intron RmInt1

High-throughput sequencing reveals circular substrates for an archaeal RNA ligase

Circular Noncoding Ribosomal Transcripts and Recombination of Retrotransposons: Two Charming Secrets Revealed by a Less-Explored Human Parasite

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