What are some alternative names used for antisense RNA?

Antisense RNA is also referred to as antisense transcript and natural antisense transcript. When referring to synthetic versions designed to target RNA, they are often called antisense oligonucleotides.

In which organisms have natural antisense RNAs been identified?

Natural antisense RNAs have been found in both prokaryotes, such as bacteria and plasmids, and eukaryotes, including plants and mammals.

How are natural antisense RNAs classified based on their length?

Natural antisense RNAs can be classified into two categories based on their length: short non-coding RNAs, which are less than 200 nucleotides long, and long non-coding RNAs, which are greater than 200 nucleotides long.

What is the main function of antisense RNA in biological systems?

The primary function of antisense RNA is to regulate gene expression. It achieves this by interacting with specific messenger RNA molecules, preventing them from being translated into proteins.

What are the applications of synthetically produced antisense RNAs?

Synthetically produced antisense RNAs are widely used as research tools for gene knockdown, a process that reduces or silences the expression of specific genes. They also hold potential for therapeutic applications.

Describe the discovery of micF asRNA in Escherichia coli.

The micF asRNA was discovered during research on the outer membrane porin protein OmpC in Escherichia coli. Clones related to the OmpC promoter were found to repress the expression of another porin, OmpF. The region responsible was a 300 base-pair locus upstream of the OmpC promoter, which showed significant sequence homology (70%) with the 5' end of the OmpF mRNA, indicating it was an antisense transcript capable of downregulating OmpF expression under stress conditions by inducing mRNA degradation.

How are most antisense RNAs identified today, beyond accidental discoveries like micF?

Beyond accidental discoveries, the majority of antisense RNAs are identified through genome-wide searches for small regulatory RNAs and comprehensive transcriptome analysis.

What computational approaches are used to predict antisense RNAs?

Computational searches for antisense RNAs typically involve identifying regions that are predicted to have conserved RNA structures, act as orphan promoters, and possess Rho-independent terminators, while excluding known protein-encoding regions.

What is a limitation of computational searches for identifying antisense RNAs?

A limitation of computational searches is that they often focus on intergenic regions, potentially missing antisense RNAs that are transcribed from the opposite strand of an encoding gene.

How can antisense RNAs transcribed from the same region as encoding genes be detected?

Antisense RNAs transcribed from the same region as encoding genes can be detected using methods like oligonucleotide microarrays, where one or both strands of the encoding genes are used as probes.

What significant discovery in 1978 laid the groundwork for antisense RNA as a therapeutic concept?

In 1978, Paul Zamecnik and Stephenson discovered an antisense oligonucleotide that was complementary to the viral RNA of Rous sarcoma virus. This oligonucleotide demonstrated the capability to inhibit viral replication and protein synthesis, establishing the potential of antisense molecules as therapeutic agents.

What was the first FDA-approved antisense RNA drug, and what was its purpose?

The first FDA-approved antisense RNA drug was fomivirsen, approved in 1998. It was a 21-base-pair oligonucleotide designed to treat cytomegalovirus retinitis in patients with AIDS by targeting and inhibiting the replication of the cytomegalovirus.

Although fomivirsen was discontinued, what was its significance in the field of antisense RNA?

Despite being discontinued in 2004 due to market reasons, fomivirsen served as a crucial successful example, demonstrating the viability of using antisense RNAs as drug targets and candidates for treating diseases.

What is mipomersen, and what is its therapeutic application?

Mipomersen is an antisense oligonucleotide approved by the FDA in 2013. It is used to manage low-density lipoprotein cholesterol (LDL) levels in patients with homozygous familial hypercholesterolemia (HoFH), a rare genetic condition characterized by extremely high cholesterol levels.

How does mipomersen work to reduce LDL cholesterol levels?

Mipomersen works by binding to the messenger RNA (mRNA) that codes for apolipoprotein B-100 (apo-B-100). Apo-B-100 is essential for producing very low-density lipoprotein (VLDL) and LDL. By targeting this mRNA for degradation via RNAse H, mipomersen reduces the production of apo-B-100, ultimately lowering LDL levels.

Where were some of the earliest antisense RNAs discovered, and what roles did they play?

Some of the earliest antisense RNAs were discovered in prokaryotes, including plasmids, bacteriophages, and bacteria. For example, in the plasmid ColE1, an asRNA called RNA I regulates the plasmid's copy number by controlling replication, and in bacteriophage P22, the asRNA sar helps regulate the switch between lytic and lysogenic cycles.

Explain the function of RNA I in the ColE1 plasmid.

In the ColE1 plasmid, RNA I acts as an antisense RNA that controls the plasmid's copy number. It does this by forming a duplex with RNA II, which is a primer RNA necessary for replication. This duplex formation causes a conformational change in RNA II, preventing it from hybridizing with its DNA template and thus resulting in a low copy number of the plasmid.

How do antisense RNAs regulate gene expression in plants, using the FLC gene as an example?

In plants like Arabidopsis thaliana, antisense RNAs regulate gene expression epigenetically. For instance, the antisense RNA of the Flowering Locus C (FLC) gene, known as COOLAIR, is expressed in cold environments. COOLAIR inhibits FLC gene expression through chromatin modification, which in turn allows for flowering to occur.

What is the role of the XIST antisense RNA in mammalian cells?

In mammalian cells, the XIST antisense RNA plays a crucial role in X chromosome inactivation. XIST recruits the polycomb repressive complex 2 (PRC2), which leads to the heterochromatinization of one of the X chromosomes, effectively silencing it.

What are the main ways antisense RNAs can be classified?

Antisense RNAs can be classified in several ways, including by their regulatory mechanisms (e.g., RNA-DNA, RNA-RNA, RNA-protein interactions), the type of promoters initiating their expression, their length (short vs. long non-coding RNAs), by species, and importantly, by their genomic location relative to their target genes as either cis-acting or trans-acting.

What defines a cis-acting antisense RNA?

A cis-acting antisense RNA is transcribed from the opposite strand of a target gene at the same genetic locus. These asRNAs typically exhibit a high degree of complementarity with their target gene.

How do cis-acting antisense RNAs regulate gene expression at the post-transcriptional level?

Cis-acting antisense RNAs that target messenger RNAs can function by either blocking the ribosome's ability to bind to the mRNA, thereby preventing translation, or by recruiting enzymes like RNAase H to degrade the target mRNA. Both mechanisms ultimately repress the translation of the targeted mRNA.

How do cis-acting epigenetic regulators differ from cis-acting asRNAs that target mRNAs?

Unlike cis-acting asRNAs that target individual mRNAs for translational repression or degradation, cis-acting epigenetic regulators function by recruiting chromatin-modifying enzymes. These enzymes can alter the epigenetic state around the locus where the asRNA is transcribed, influencing both the transcription locus itself and neighboring genes.

What characterizes trans-acting antisense RNAs in terms of their location and properties?

Trans-acting antisense RNAs are transcribed from loci that are located at a distance from their target genes. In contrast to cis-acting asRNAs, they generally display a lower degree of complementarity with their targets, can be longer, and may target multiple genes.

What factors contribute to the complexity and potential challenges in targeting trans-acting antisense RNAs?

Trans-acting antisense RNAs form less stable complexes with their target transcripts due to lower complementarity. They may also require the assistance of RNA chaperone proteins, like Hfq, to function effectively. These complexities currently make them considered less viable targets for drug development compared to cis-acting asRNAs.

What are the primary mechanisms by which antisense RNAs regulate gene expression?

Antisense RNAs regulate gene expression through three main mechanisms: epigenetic regulation (involving DNA methylation and histone modification), co-transcriptional regulation (affecting transcription initiation or elongation), and post-transcriptional regulation (directly targeting mRNAs).

How can antisense RNAs induce DNA methylation to downregulate gene expression?

Some antisense RNAs can recruit DNA methyltransferases to specific gene promoters. This recruitment leads to DNA methylation, a process that can result in the long-term silencing or downregulation of the targeted genes.

Provide an example of an antisense RNA involved in DNA methylation and disease.

The antisense non-coding RNA in the INK locus (ANRIL) is an example. It is expressed in the same locus as the tumor suppressor gene p15INK4b and is responsible for silencing this gene in certain types of leukemia by inducing DNA methylation of its promoter.

How do antisense RNAs influence gene expression through histone modification?

Antisense RNAs can recruit chromatin-modifying enzymes, such as polycomb repressive complex 2 (PRC2), which are responsible for histone modifications like histone methylation. These modifications can alter chromatin structure and lead to gene repression.

Explain the role of ANRIL in histone modification.

ANRIL, in addition to inducing DNA methylation, can also repress the neighboring gene CDKN2A by recruiting polycomb repressive complex 2 (PRC2). This recruitment leads to histone methylation (specifically H3K27me) at the CDKN2A locus, contributing to gene silencing.

How can antisense RNAs regulate gene expression during the transcription process (co-transcriptional regulation)?

Antisense RNAs can affect gene expression co-transcriptionally in several ways. They can cause RNA polymerase collisions, leading to premature transcription termination. They can also cause polymerase pausing, slowing down or halting transcription elongation. Additionally, they can influence splicing or maintain specific splice variants, thereby affecting the final mRNA product.

How does the ZEB2 asRNA impact the synthesis of E-cadherin?

The ZEB2 asRNA influences the synthesis of E-cadherin by maintaining an internal ribosome entry site (IRES) within the ZEB2 mRNA. This IRES is crucial for efficient translation of ZEB2 mRNA into the protein E-cadherin, which acts as a transcriptional repressor.

What is post-transcriptional regulation by antisense RNA?

Post-transcriptional regulation by antisense RNA involves the direct targeting of messenger RNAs (mRNAs) by asRNAs. This interaction affects the translation of the mRNA into protein, either by blocking ribosome binding or by promoting mRNA degradation.

What are the advantages of using antisense RNAs as therapeutic targets?

Antisense RNAs offer several advantages as therapeutic targets: they regulate gene expression at multiple levels (transcription, post-transcription, epigenetics), cis-acting asRNAs are highly sequence-specific, and their typically low expression levels mean only small dosages are needed for therapeutic effect.

What is the challenge in developing drugs that increase gene expression, and how might antisense RNAs help?

Developing drugs that upregulate gene expression is generally more challenging than creating inhibitors. However, by designing molecules called antagoNATs to inhibit specific asRNAs, researchers aim to release the repression these asRNAs impose, thereby increasing the expression of targeted genes, such as tumor suppressor genes.

What are antagoNATs, and what is their therapeutic goal?

AntagoNATs (antisense oligonucleotides targeting natural antisense transcripts) are single-stranded oligonucleotides designed to inhibit the function of endogenous antisense RNAs. Their therapeutic goal is to restore or increase the expression of specific genes that are normally downregulated by these asRNAs.

What are the main challenges associated with using antisense RNAs and antagoNATs as therapeutics?

Key challenges include the potential for degradation by enzymes like RNase, requiring chemical modifications (like phosphorothioate linkages) which can cause side effects; off-target toxicity due to imperfect targeting; and limited intracellular uptake, necessitating the use of carriers for efficient delivery.

What is a common chemical modification used for therapeutic oligonucleotides, and what are its drawbacks?

A common chemical modification is adding a phosphorothioate linkage to the backbone of the oligonucleotide to prevent degradation. However, this modification can be proinflammatory and may cause adverse effects such as fever, chills, or nausea.

How does off-target toxicity manifest with synthesized antisense oligonucleotides?

Off-target toxicity occurs when synthesized oligonucleotides bind to unintended targets. This can happen because even a single mismatch between the oligonucleotide and its target sequence can distort the structure required for proper recognition and function, leading to unexpected effects.

What are the limitations regarding the intracellular uptake of artificial antisense RNAs?

Artificial antisense RNAs generally have limited ability to enter cells on their own. While some cell types like neurons and glia can uptake naked oligonucleotides, using traceable carriers, such as viral vectors or lipid vesicles, is often preferred to control and monitor their concentration and metabolism within the cell.

How does the provided image caption describe antisense RNA?

The image caption explains that antisense RNA (asRNA) is transcribed from the lagging strand of a gene and is complementary to a specific messenger RNA (mRNA) or sense transcript.

Antisense Rna Wiki: Antisense RNA: The Unseen Regulators

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Introduction

Complementary Binding

Antisense RNA (asRNA), also known as a natural antisense transcript (NAT), is a single-stranded RNA molecule that is complementary to a protein-coding messenger RNA (mRNA). By hybridizing with its target mRNA, asRNA effectively inhibits the translation of that mRNA into protein, thereby playing a crucial role in regulating gene expression.

Ubiquitous Presence

These regulatory molecules are not confined to specific organisms; asRNAs have been identified in both prokaryotes and eukaryotes. They can be classified based on their length into short non-coding RNAs (ncRNAs) typically less than 200 nucleotides, and long non-coding RNAs (lncRNAs) exceeding 200 nucleotides.

Research and Therapy

Beyond their natural biological functions, asRNAs are invaluable tools in molecular biology research for targeted gene knockdown. Furthermore, their specific regulatory capabilities have spurred significant interest in their potential therapeutic applications.

Discovery and Development

Early Observations

The initial discovery of asRNAs often occurred serendipitously during investigations into protein functions. For instance, the micF asRNA was identified while studying the outer membrane porin OmpC in E. coli. Researchers observed that certain OmpC promoter clones could repress the expression of other porins like OmpF. The region responsible was found to be homologous to the 5' end of OmpF mRNA, acting as an asRNA that downregulates OmpF expression under stress conditions by forming a duplex with its mRNA, leading to degradation.

High-Throughput Identification

While early discoveries were often accidental, the majority of asRNAs have since been identified through comprehensive, genome-wide searches employing techniques like transcriptome analysis and oligonucleotide microarrays. Modern approaches focus on strand-specific transcription, chromatin interactions, and single-cell studies to enhance accuracy and minimize false positives.

Therapeutic Milestones

The concept of targeting asRNAs therapeutically emerged as early as 1978. The first FDA-approved asRNA-based drug was Fomivirsen in 1998, used to treat cytomegalovirus retinitis in AIDS patients by inhibiting viral replication. Although discontinued, it demonstrated the viability of asRNAs as therapeutic agents. More recently, Mipomersen received FDA approval in 2013 for managing severe hypercholesterolemia by targeting the mRNA encoding apolipoprotein B-100.

Examples Across Species

Prokaryotic Systems

Antisense RNAs were first characterized in prokaryotes. Examples include RNA I in the plasmid ColE1, which regulates plasmid copy number by controlling replication primer RNA II, and the sar asRNA in bacteriophage P22, which modulates the lytic/lysogenic cycle by controlling Ant expression.

Plant Regulation

In plants, asRNAs play roles in developmental processes. The COOLAIR asRNA regulates the Flowering Locus C (FLC) gene in Arabidopsis thaliana, influencing floral transition, particularly under cold conditions. Additionally, asDOG1 (or 1GOD) acts in cis to negatively regulate the expression of the Delay of Germination 1 (DOG1) gene.

Mammalian Mechanisms

In mammals, a prominent example is the XIST asRNA, crucial for X chromosome inactivation. XIST recruits Polycomb Repressive Complex 2 (PRC2), leading to heterochromatinization and silencing of one X chromosome in females. Another example, HOTAIR, transcribed from the HOXC locus, acts trans-acting to recruit PRC2 to the HOXD locus, causing gene silencing.

Classification Schemes

Functional Categories

Antisense RNAs can be broadly categorized by their regulatory mechanisms: interactions involving RNA-DNA, RNA-RNA (within the nucleus or cytoplasm), and RNA-protein complexes, often leading to epigenetic modifications.

Cis-acting vs. Trans-acting

A key classification is based on the genomic location relative to the target gene:

Cis-acting asRNAs: Transcribed from the opposite strand of the target gene locus. They typically exhibit high complementarity, directly targeting individual mRNAs for translational repression or degradation, or regulating epigenetic states at their own locus.
Trans-acting asRNAs: Transcribed from loci distant from the target genes. They generally show lower complementarity, can target multiple loci, and may require accessory proteins like Hfq for function. They are often considered more complex and less amenable to therapeutic targeting.

Mechanisms of Action

Epigenetic Regulation

asRNAs can profoundly influence gene expression through epigenetic modifications. They can recruit DNA methyltransferases (DNMTs) to induce DNA methylation, leading to long-term gene silencing, as seen with the HBA1 gene in alpha-thalassemia or the tumor suppressor gene p15INK4b via the ANRIL transcript. Similarly, they can recruit histone methyltransferases (HMTs) or complexes like PRC2 to induce histone methylation, altering chromatin structure and accessibility. Examples include ANRIL's role in repressing CDKN2A and XIST's role in X chromosome inactivation.

Co-transcriptional Regulation

Regulation can also occur during the transcription process itself. asRNAs can induce polymerase collision, leading to premature transcription termination. They can also cause polymerase pausing, hindering elongation. Furthermore, they can influence alternative splicing outcomes or fine-tune the expression levels of specific transcript isoforms, presenting a sophisticated layer of gene control.

Post-transcriptional Control

The most direct mechanism involves asRNAs targeting mature mRNAs. By forming duplexes, they can sterically hinder ribosome binding, thereby inhibiting translation. Alternatively, the asRNA-mRNA duplex can recruit enzymes like RNase H, leading to mRNA degradation. While inhibitory effects are more common, some asRNAs have been shown to activate translation under specific circumstances.

Therapeutic Potential & Challenges

Advantages for Drug Development

Antisense RNAs offer several advantages as therapeutic targets or agents. Their ability to regulate gene expression at multiple levels (transcription, post-transcription, epigenetics), their inherent sequence specificity (particularly cis-acting variants), and the potential for low-dose efficacy due to their regulatory nature make them attractive candidates.

A significant area of interest is developing therapies that *inhibit* asRNAs (using antagoNATs) to restore or enhance the expression of beneficial genes, such as tumor suppressors or genes silenced in genetic disorders.

Overcoming Hurdles

Despite their promise, significant challenges remain. The inherent instability of RNA necessitates chemical modifications (e.g., phosphorothioate linkages) to prevent degradation, though these can sometimes cause proinflammatory side effects. Off-target toxicity is another concern, as even minor sequence mismatches can disrupt binding and lead to unintended effects. Furthermore, achieving efficient and controlled intracellular uptake of synthetic antisense oligonucleotides remains an area of active research, often requiring specialized delivery vehicles.

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References

Weiss, B. (ed.): Antisense Oligodeoxynucleotides and Antisense RNA : Novel Pharmacological and Therapeutic Agents, CRC Press, Boca Raton, FL, 1997.

A full list of references for this article are available at the Antisense RNA Wikipedia page

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This page was generated by an Artificial Intelligence and is intended for informational and educational purposes only. The content is based on a snapshot of publicly available data from Wikipedia and may not be entirely accurate, complete, or up-to-date.

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Antisense RNA: The Unseen Regulators

💡 Dive in with Flashcard Learning! 💡

Introduction ℹ️

🔗 Complementary Binding

🌐 Ubiquitous Presence

🔬 Research and Therapy

Discovery and Development 📜

💡 Early Observations

📊 High-Throughput Identification

💊 Therapeutic Milestones

Examples Across Species 🌳

🦠 Prokaryotic Systems

🌿 Plant Regulation

🐾 Mammalian Mechanisms

Classification Schemes 🗂️

⚙️ Functional Categories

📍 Cis-acting vs. Trans-acting

Mechanisms of Action 🛠️

epigenetic Epigenetic Regulation

🔄 Co-transcriptional Regulation

📉 Post-transcriptional Control

Therapeutic Potential & Challenges 🧑‍⚕️

✅ Advantages for Drug Development

⚠️ Overcoming Hurdles

Teacher's Corner 🧑‍🏫

Edit and Print this course in the Wiki2Web Teacher Studio

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References

📜 References

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Disclaimer ⚠️

📜 Important Notice

Dive in with Flashcard Learning!

Introduction

Complementary Binding

Ubiquitous Presence

Research and Therapy

Discovery and Development

Early Observations

High-Throughput Identification

Therapeutic Milestones

Examples Across Species

Prokaryotic Systems

Plant Regulation

Mammalian Mechanisms

Classification Schemes

Functional Categories

Cis-acting vs. Trans-acting

Mechanisms of Action

Epigenetic Regulation

Co-transcriptional Regulation

Post-transcriptional Control

Therapeutic Potential & Challenges

Advantages for Drug Development

Overcoming Hurdles

Teacher's Corner

True or False?

Test Your Knowledge!

Gamer's Corner

Discover other topics to study!

References

Feedback & Support

Disclaimer

Important Notice