Detecting of chloroplast circular RNAs in Arabidopsis thaliana.
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ABSTRACT:
SUBMITTER: Liu S
PROVIDER: S-EPMC6619943 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Circular RNAs (circRNAs) are a family of transcripts with covalently closed circular structures and still largely unknown functions. Large numbers of circRNAs have been found in various biological processes in humans and animals, but fewer circRNAs have been identified in plants. We performed a genome-wide analysis of circRNAs in Arabidopsis thaliana via deep sequencing. We constructed 14 strand-specific libraries from 13 samples of plants from four developmental stages, four stress treatments, and five organs and a mixed sample across the lifespan. In total, we identified 5861 circRNAs, including 1275 novel ones, using the strict threshold of at least two unique back-spliced supporting reads. The circRNAs were non-randomly distributed in all chromosomes; most were exonic. Sequence similarity analysis of circRNAs between A. thaliana and four other species showed that some circRNAs are conserved in plants. Functional annotation indicated that many parent genes of circRNAs are involved in many fundamental processes including plant development, reproduction, and response to stimulus. In addition, a small proportion of circRNAs was shown to be potential targets of miRNAs, indicating that the circRNAs could interact with miRNAs to regulate gene expression. qRT-PCR analysis revealed that circRNAs displayed diverse expression patterns at different growth stages. Our results provide an important resource for continuing circRNA research in A. thaliana, and should enhance our understanding of circRNAs in plants.
| S-EPMC5623955 | biostudies-literature
Project description:The pervasive expression of circular RNAs (circRNAs) is a recently discovered feature of gene expression in highly diverged eukaryotes. Numerous algorithms that are used to detect genome-wide circRNA expression from RNA sequencing (RNA-seq) data have been developed in the past few years, but there is little overlap in their predictions and no clear gold-standard method to assess the accuracy of these algorithms. We review sources of experimental and bioinformatic biases that complicate the accurate discovery of circRNAs and discuss statistical approaches to address these biases. We conclude with a discussion of the current experimental progress on the topic.
| S-EPMC5565156 | biostudies-other
Project description:BACKGROUND:Recently, identification and functional studies of circular RNAs, a type of non-coding RNAs arising from a ligation of 3' and 5' ends of a linear RNA molecule, were conducted in mammalian cells with the development of RNA-seq technology. METHOD:Since compared with animals, studies on circular RNAs in plants are less thorough, a genome-wide identification of circular RNA candidates in Arabidopsis was conducted with our own developed bioinformatics tool to several existing RNA-seq datasets specifically for non-coding RNAs. RESULTS:A total of 164 circular RNA candidates were identified from RNA-seq data, and 4 circular RNA transcripts, including both exonic and intronic circular RNAs, were experimentally validated. Interestingly, our results show that circular RNA transcripts are enriched in the photosynthesis system for the leaf tissue and correlated to the higher expression levels of their parent genes. Sixteen out of all 40 genes that have circular RNA candidates are related to the photosynthesis system, and out of the total 146 exonic circular RNA candidates, 63 are found in chloroplast.
| S-EPMC5635619 | biostudies-literature
Project description:Competing endogenous RNAs (ceRNAs) are natural transcripts that can act as endogenous sponges of microRNAs (miRNAs), modulating miRNA action upon target mRNAs. Circular RNAs (circRNAs) are one among the various classes of ceRNAs. They are produced from a process called back-splicing and have been identified in many eukaryotes. In plants, their effective action as a miRNA sponge was not yet demonstrated. To address this question, public mRNAseq data from Argonaute-immunoprecipitation libraries (AGO-IP) of Arabidopsis thaliana flowers were used in association with a bioinformatics comparative multi-method to identify putative circular RNAs. A total of 27,812 circRNAs, with at least two reads at the back-splicing junction, were identified. Further analyses were used to select those circRNAs with potential miRNAs binding sites. As AGO forms a ternary complex with miRNA and target mRNA, targets count in AGO-IP and input libraries were compared, demonstrating that mRNA targets of these miRNAs are enriched in AGO-IP libraries. Through this work, five circRNAs that may function as miRNA sponges were identified and one of them were validated by PCR and sequencing. Our findings indicate that this post-transcriptional regulation can also occur in plants.
| S-EPMC6784238 | biostudies-literature
Project description:In all living organisms, regulation of gene expression is fundamental for survival and adaptation. Gene expression can be modulated at various steps, including at the level of RNA processing. During the last few years, the importance of alternative splicing of mRNAs in controlling plant development and stress responses were emerged and highlighted its importance. Recently, an other type of alternative splicing has been reported which leads to the generation of circular RNAs (circRNAs), a novel class on endogenous noncoding RNAs. Several functions of circular RNAs have been proven or proposed, including functioning as microRNA or RNA-binding protein decoys, playing regulatory roles in gene expression or affecting transcriptional control via special RNA-RNA interactions. Despite the widening knowledge of circRNAs and their functional aspects in the animal kingdom, relatively little is known about circRNAs in plants. In order to detect and classify circRNAs in Arabidopsis thaliana, we created a workflow that includes generation of Illumina libraries enriched for circRNAs and a comparison of biocomputational tools developed for detecting endogenous circular RNAs in other species. With the power of high-throughput sequencing and evaluation of algorithms, high-fidelity candidates were subjected for an analysis of their functional role in plant development and stress-related responses, especially regarding the role of splicing, including alternative splicing events, splice site preference and strength variances and transcript composition and to comprehend the role of RNA processing in stress response. Here we present an approach combining bioinformatic tools and molecular techniques to investigate the adaptability of detection methods of circRNAs from other species to plant circular RNAs, and based on our high-fidelity results identify and understand the characteristics of circRNAs in Arabidopsis thaliana.
2019-01-24 | GSE125378 | GEO
Project description:To exlore more circRNAs involved in Arabidopsis thaliana, we deeply sequenced 14 samples including whole plants from four developmental stages (rosette leaves > 1 mm in length; rosette growth complete; 50% of flowers to be produced have opened; first silique shattered), aerial part of plants from four stress treatments (control, drought, salinity and heat), five organs (roots, stems, leaves, flowers and siliques) and a mixed sample from whole plants across the lifespan (cotyledons emergence, rosette leaves﹥1 mm, rosette growth complete, first flower open, flourishing florescence, first silique shattered, senescence). The total RNA was purified by rRNA-depletion and linear RNA removal with RNAseR, and paired-end (PE) sequenced by Illumina HiSeq 2500 (read length, PE125, the mixed sample) and Illumina Hiseq X Ten (read length, PE150, 13 independent samples) platforms. We obtained 181.97 Gb raw data (151.37 Gb from 13 samples and 30.6 Gb from a mixed sample) and identified 5861 circRNAs with expression quantity. We annotated the parent genes of these circRNAs and predicted their target sites of microRNAs.
2016-09-01 | GSE77672 | GEO
Project description:Whole-genome oligonucleotide resequencing arrays have allowed the comprehensive discovery of single nucleotide polymorphisms (SNPs) in eukaryotic genomes of moderate to large size. With this technology, the detection rate for isolated SNPs is typically high. However, it is greatly reduced when other polymorphisms are located near a SNP as multiple mismatches inhibit hybridization to arrayed oligonucleotides. Contiguous tracts of suppressed hybridization therefore typify polymorphic regions (PRs) such as clusters of SNPs or deletions. We developed a machine learning method, designated margin-based prediction of polymorphic regions (mPPR), to predict PRs from resequencing array data. Conceptually similar to hidden Markov models, the method is trained with discriminative learning techniques related to support vector machines, and accurately identifies even very short polymorphic tracts (<10 bp). We applied this method to resequencing array data previously generated for the euchromatic genomes of 20 strains (accessions) of the best-characterized plant, Arabidopsis thaliana. Nonredundantly, 27% of the genome was included within the boundaries of PRs predicted at high specificity ( approximately 97%). The resulting data set provides a fine-scale view of polymorphic sequences in A. thaliana; patterns of polymorphism not apparent in SNP data were readily detected, especially for noncoding regions. Our predictions provide a valuable resource for evolutionary genetic and functional studies in A. thaliana, and our method is applicable to similar data sets in other species. More broadly, our computational approach can be applied to other segmentation tasks related to the analysis of genomic variation.
| S-EPMC2413159 | biostudies-literature
Project description:Circular RNAs (circRNAs) are the products of the non-canonical splicing of pre-mRNAs. In contrast to humans and animals, our knowledge of the biogenesis and function of circRNAs in plants is very scarce. To identify proteins involved in plant circRNA generation, we characterized the transcriptomes of 18 Arabidopsis thaliana knockout mutants for genes related to splicing. The vast majority (>90%) of circRNAs were formed in more than one variant; only a small fraction of circRNAs was mutant-specific. Five times more circRNA types were identified in cbp80 and three times more in c2h2 mutants than in the wild-type. We also discovered that in cbp80, c2h2 and flk mutants, the accumulation of circRNAs was significantly increased. The increased accumulation of circular transcripts was not accompanied by corresponding changes in the accumulation of linear transcripts. Our results indicate that one of the roles of CBP80, C2H2 and FLK in splicing is to ensure the proper order of the exons. In the absence of one of the above-mentioned factors, the process might be altered, leading to the production of circular transcripts. This suggests that the transition toward circRNA production can be triggered by factors sequestering these proteins. Consequently, the expression of linear transcripts might be regulated through circRNA production.
| S-EPMC7564263 | biostudies-literature
Project description:Arabidopsis thaliana chloroplasts contain at least two 3' to 5' exoribonucleases, polynucleotide phosphorylase (PNPase) and an RNase R homolog (RNR1). PNPase has been implicated in both mRNA and 23S rRNA 3' processing. However, the observed maturation defects do not affect chloroplast translation, suggesting that the overall role of PNPase in maturation of chloroplast rRNA is not essential. Here, we show that this role can be largely ascribed to RNR1, for which homozygous mutants germinate only on sucrose-containing media, and have white cotyledons and pale green rosette leaves. Accumulation of chloroplast-encoded mRNAs and tRNAs is unaffected in such mutants, suggesting that RNR1 activity is either unnecessary or redundant for their processing and turnover. However, accumulation of several chloroplast rRNA species is severely affected. High-resolution RNA gel blot analysis, and mapping of 5' and 3' ends, revealed that RNR1 is involved in the maturation of 23S, 16S and 5S rRNAs. The 3' extensions of the accumulating 5S rRNA precursors can be efficiently removed in vitro by purified RNR1, consistent with this view. Our data suggest that decreased accumulation of mature chloroplast ribosomal RNAs leads to a reduction in the number of translating ribosomes, ultimately compromising chloroplast protein abundance and thus plant growth and development.
| S-EPMC1110743 | biostudies-literature
Project description:AtRsmD was recently demonstrated to be a chloroplast 16S rRNA methyltransferase (MTase) for the m2G915 modification in Arabidopsis. Here, its function of AtRsmD for chloroplast development and photosynthesis was further analyzed. The AtRsmD gene is highly expressed in green photosynthetic tissues. AtRsmD is associated with the thylakoid in chloroplasts. The atrsmd-2 mutant exhibited impaired photosynthetic efficiency in emerging leaves under normal growth conditions. A few thylakoid lamellas could be observed in the chloroplast from the atrsmd-2 mutant, and these thylakoids were loosely organized. Knockout of the AtRsmD gene had minor effects on chloroplast ribosome biogenesis and RNA loading on chloroplast ribosomes, but it reduced the amounts of chloroplast-encoded photosynthesis-related proteins in the emerging leaves, for example, D1, D2, CP43, and CP47, which reduced the accumulation of the photosynthetic complex. Nevertheless, knockout of the AtRsmD gene did not cause a general reduction in chloroplast-encoded proteins in Arabidopsis grown under normal growth conditions. Additionally, the atrsmd-2 mutant exhibited more sensitivity to lincomycin, which specifically inhibits the elongation of nascent polypeptide chains. Cold stress exacerbated the effect on chloroplast ribosome biogenesis in the atrsmd-2 mutant. All these data suggest that the AtRsmD protein plays distinct regulatory roles in chloroplast translation, which is required for chloroplast development and chloroplast function.
| S-EPMC9083416 | biostudies-literature