The ribo-circRNA button features three dropdown options, which provide access to content pages: ① Computationally predicted ribo-circRNAs ② Literature-reported trans-circRNAs ③ Cross-species conserved ribo-circRNAs

1. Computationally predicted ribo-circRNAs
Each ribo-circRNAs is characterized using ribosome profiling data and presented at two distinct levels: ① Condition-independent prediction: Based on previously documented circRNAs and Ribo-seq data, and ② Condition-dependent prediction: Derived from public RNA-seq and matched Ribo-seq data. Characterized ribo-circRNAs come with meta-information such as riboCIRC_ID (self-named according to "a guide to naming human non-coding RNA genes"), circRNA_coordinates (chr:start-end), Host_gene (Ensembl_ID, Symbol, and Transcript), and circRNA length (nt). Upon clicking the riboCIRC_ID or the More details button, users can access detailed information, including structure and designed circRNA primer sets.

2. Literature-reported trans-circRNAs
This page features a manually curated encyclopedia of experimentally validated circRNAs capable of peptide generation. Detailed meta-information for each circRNA is provided, including documented circRNA_Name, Mapped CircBase_ID, circRNA_coordinates (chr:start-end), Strand, HostGene (Transcript), circRNA derived ORF (Length(aa) and peptide sequence), Experimental evidence (MS data supporting, Exogenous expression, and Antibody detection of endogenous peptide), and related reference (PubMed_ID, Paper, Journal and Release year).

3. Cross-Species Conserved ribo-circRNAs
This page displays translatable circRNAs conserved across species, with conserved entries aligned in the same row for easy comparison.

The Circ-peptide page provides a systematic de novo sequence, structural, and functional annotation on circRNA-encoded peptides based on bioinformatics prediction. When clicking on the items enclosed in the container box to select one from the dropdown list, more information about circRNA-encoded peptides will be returned, including circRNA-derived ORF annotation (cORF_coordinates (chr:start-end), cORF length(aa) and cORF_seq), cORF conservation (phastCons score), predicted peptide characteristics (signal cleavage site (SignalP), N-terminal presequences (TargetP), transmembrane domain prediction (TMHMM and Phobius), and localization (DeepLoc)), and protein structure prediction (I-TASSER). These results provide a foundation for circRNA translation studies, serving as a starting point for further investigation into their biological significance of circRNA-encoded peptides.

The visualization page provides intuitive visualization-oriented information about circRNA translation, view of junction-spanning ribosome footprints. The context-specific selection for translatabe circRNAs and ribosome profiling data tracks are accessible in the left side bar of JBrowse.

All data within this database is freely available for download. Users are welcome to utilize this data for personal, non-profit, and academic purposes without restriction.

This page provides a grand summary that aggregates data from all records currently visible in the database.

The Feedback page offers a dedicated form where you can submit comments on published ribo-circRNAs or ribosome profiling data. We also welcome any specific and helpful suggestions you might have—please don't hesitate to contact us.

1. Ribosome Profiling Characterization of circRNAs

Two distinct strategies were here used to characterize ribo-circRNAs: (1) Condition-independent prediction based on previously documented circRNAs and Ribo-seq data and (2) Condition-dependent prediction derived from public RNA-seq and their matched Ribo-seq data.

We employed CIRCexplorer3 and find_circ to identify transcribed circRNAs within each RNA-seq dataset. For full-length circRNA sequences, we reconstructed them using the circRNAFull package or extracted them from databases like circAtlas, CSCD, circRNADb, circBank, CIRCpedia, and circBase if RNA-seq data wasn't available. To pinpoint ribosome-associated circRNAs, we first mapped all ribosome-protected footprints to the corresponding linear reference genome using Tophat2. Subsequently, the resulting unmapped.bam files were remapped to our pseudo circRNA reference using BWA. Finally, a circRNA was designated as associated with translating ribosomes only if it simultaneously met these four criteria: (1) At least two unique backsplice junction-spanning footprints, (2) A minimum read-junction overlap of three nucleotides (nt) on either side of the backsplice junction site, (3) A Ribo-seq read length range of 25-35 nt, and (4) Possession of a BSJ-spanning ORF with an NTG start codon.

2. Detection of Cross-Species Conserved ribo-circRNAs

The process is detailed below: First, candidate orthologous genes are obtained from the OMA Orthologs Database. Then, 50 bp sequences flanking the circRNA backsplice junction (BSJ) are extracted to represent the BSJ sequence (if the length is less than 100 bp, the sequence is clipped from the center of the circRNA). Subsequently, all ribo-circRNA BSJ sequences from one species are aligned against those from other species using BLAT, with parameters set to -minIdentity=30 -tileSize=6. We then filter and retain alignment entries that are both orthologous genes and show an identity exceeding 50%. Finally, the pair with the highest mutual alignment identity is selected as the definitive orthologous ribo-circRNA pair.