As research progresses, circular RNAs (circRNAs, also known as cyclic RNAs) have the potential to revolutionize diagnostics, therapeutics, and biotechnology. Small circRNAs are non-coding RNAs characterized by their covalently closed-loop structure, which makes them resistant to exonucleases. Unlike linear RNAs, they lack free 5'- and 3'-ends. CircRNAs are generated primarily through back-splicing, where a downstream splice donor site joins with an upstream splice acceptor site within pre-mRNA. CircRNAs have emerged as critical players in cellular biology, with diverse applications and roles. Their unique properties, such as stability, abundance, and tissue-specific expression, make them intriguing molecules for both basic research and translational applications. Small Circular RNAs are typically less than 1,000 nucleotides long and are a subset of circRNAs that can vary significantly in size. Their circular nature enhances their stability in cells compared to linear RNAs.
Structures of circular or cyclic adenosines, cA4 and cA6
Known roles of circRNAs
miRNA sponges: Some circRNAs can bind and sequester microRNAs, preventing them from interacting with their mRNA targets.
Protein interactions: CircRNAs may interact with RNA-binding proteins, influencing their activity.
Translation: Although most circRNAs are non-coding, some small circRNAs can code for proteins under specific conditions.
Biogenesis: CircRNAs arise from precursor mRNA (pre-mRNA) during splicing, involving the canonical spliceosomal machinery, but in a non-linear arrangement.
Type III CRISPR system: Cyclic oligoadenylate (cOA) molecules are second messengers in the type III CRISPR system. cOAs activate promiscuous ancillary nucleases that indiscriminately degrade host and viral DNA/RNA. Type III CRISPR-Cas systems provide adaptive immunity against foreign mobile genetic elements through RNA-guided interference. Sequence-specific recognition of RNA targets by the type III effector complex triggers the generation of cOA second messengers that activate ancillary effector proteins, reinforcing the host immune response. Cyclic tetra-AMP (cA4) activates the ancillary nuclease Can2.
Biological Relevance
CircRNAs are involved in diverse cellular processes, expressed in a tissue- and developmental-stage-specific manner. Dysregulated circRNAs have been linked to diseases such as cancer, neurological disorders, and cardiovascular diseases.
Roles of circRNAs in Cellular Biology
MicroRNA (miRNA) Sponges: CircRNAs can act as competitive endogenous RNAs (ceRNAs), binding to miRNAs and preventing them from interacting with their mRNA targets, for example, CDR1as (ciRS-7) with multiple binding sites for miR-7, modulating miR-7's downstream effects.
Protein Interaction Scaffolds: CircRNAs can bind and modulate the activity of RNA-binding proteins (RBPs), acting as molecular sponges or scaffolds influencing processes like transcription, splicing, or protein localization.
Translation into Functional Peptides: Some circRNAs contain internal ribosome entry sites (IRESs) or N6-methyladenosine (m6A) modifications, enabling translation into proteins or peptides. These peptides can have regulatory or functional roles in cells.
Regulation of Gene Expression: CircRNAs can influence gene transcription or post-transcriptional processing by interacting with the transcriptional machinery or splicing factors.
Epigenetic Modulation: Emerging evidence suggests circRNAs play roles in modifying chromatin structure or function indirectly through miRNA and RBP interactions.
Applications of circRNAs
Biomarkers for Disease Diagnosis
CircRNAs' stability in body fluids, such as blood, saliva, and cerebrospinal fluid, makes them ideal non-invasive disease biomarkers.
In Cancer: Altered circRNA expression profiles have been linked to tumorigenesis and metastasis. circRNAs are associated with bladder cancer and hepatocellular carcinoma occurrence and development.
In Neurological Disorders: Specific circRNAs are dysregulated in diseases like Alzheimer's and Parkinson's.
In Cardiovascular Diseases: CircRNAs play roles in heart development, myocardial infarction, and atherosclerosis.
Therapeutic Targets
Targeting of circRNAs may restore normal cellular function.
miRNA Inhibition: Blocking oncogenic circRNAs can prevent them from acting as miRNA sponges.
Gene Editing: CRISPR/Cas systems enable disruption or modification of circRNAs.
Drug Delivery Vehicles: Correctly engineered circRNAs could allow the delivery of RNA-based therapeutics, such as siRNAs, antisense oligonucleotides, or ribozymes. Their circular structure provides a stable platform for extended activity.
Protein and Peptide Therapeutics: Synthetic circRNAs encoding therapeutic peptides could allow targeting diseases where protein supplementation or functional peptides are needed.
Synthetic Biology and Biotechnology: CircRNAs may allow the design of stable RNA-based systems for gene expression in synthetic biology or as molecular scaffolds in various biotechnological applications.
Challenges and Future Directions
Biogenesis Control: Engineering circRNAs for specific applications requires precise control over their formation and properties.
Delivery Systems: Efficient methods to introduce circRNAs into cells or tissues are still needed and under development.
Functional Annotation: Many circRNAs remain uncharacterized and have unknown biological relevance.
Safety and Immunogenicity: CircRNA-based therapies must avoid triggering immune responses.
Synthesis of modified circRNA
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| Single-stranded circular DNA (circDNA) or circular RNA (circRNA) oligo-nucleotides are designed to undergo triplex formation with single-stranded (ss) DNA and RNA targets. Many of these circular oligonucleotides bind with greater affinity, sequence selectivity and are more resistant to nucleases compared to their linear counterparts. Synthetic circular DNA or circular RNA oligonucleotides can be chemically synthesized at Bio-Synthesis using solution and solid-phase synthesis methods from partially protected linear precursors. Modified nucleic acid can be incorporated chemically during synthesis as well. After removing the linear DNA, circular oligonucleotides are purified by denaturing polyacrylamide gel electrophoresis (PAGE) with >95% purity. Circularity is determined by exonuclease cleavage. As a final step, we apply an independent QA procedure to guarantee the highest quality for each circular oligonucleotide. Specs: - Length- 5to 20 mers
- Quantities- 100, 200, 500 ,1000 pmole
- Purification HPLC or PAGE purified
- Validation- MS and PAGE analysis plus additional QA procedures
For additional information regarding custom circular DNA or RNA synthesis, contact us or send an email to info@biosyn.com |
References
https://www.biosyn.com/circular-oligonucleotide.aspx
https://www.biosyn.com/tew/Synthetic-long-single-stranded-and-circular-DNA.aspx
https://www.biosyn.com/tew/what-are-circular-oligonucleotides.aspx
https://www.biosyn.com/faq/How-stable-are-circular-oligonucleotides.aspx
https://www.biosyn.com/tew/enzymatic-synthesis-of-circular-oligonucleotides.aspx
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Bio-Synthesis provides a full spectrum of high quality custom oligonucleotide modification services including 5'-triphosphate and back-bone modifications, conjugation to fatty acids, biotinylation by direct solid-phase chemical synthesis or enzyme-assisted approaches to obtain artificially modified oligonucleotides, such as BNA antisense oligonucleotides, mRNAs or siRNAs, containing a natural or modified backbone, as well as base, sugar and internucleotide linkages.
Bio-Synthesis also provides biotinylated mRNA and long circular oligonucleotides".
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