Photoactivatable A-to-I RNA Editor Advances Gene Therapy

In the rapidly evolving landscape of gene therapy, achieving precise, controllable, and reversible modulation of gene expression has long been a paramount challenge. Traditional chemical inducers deployed for controlling gene expression are often hampered by intrinsic limitations such as rapid diffusion, prolonged metabolism, and unintended systemic side effects. Addressing these critical bottlenecks, a groundbreaking study has unveiled a novel photoactivatable RNA adenosine base editor (PA-rABE), a cutting-edge technology that promises to redefine the paradigms of gene editing with unparalleled spatiotemporal control.

At the heart of this innovation lies a cleverly engineered molecular complex that integrates a compact Cas13 variant—an RNA-targeting endonuclease—with a split form of ADAR2 deaminase, the enzyme responsible for catalyzing adenosine-to-inosine (A-to-I) conversions in RNA. This fusion is further sophisticated by the incorporation of the “Magnets” system, a pair of photodimerizable protein domains that respond to blue light stimulation. Upon illumination, these Magnet domains undergo rapid and reversible dimerization, reuniting the enzymatic fragments into a fully functional base editor capable of selective RNA editing. This optical switch confers exquisite temporal precision and spatial specificity, overcoming the diffusion and metabolism concerns inherent to chemical methods.

Experimental evaluation demonstrated that PA-rABE efficiently edits endogenous RNA targets with remarkable precision. Notably, the system achieves minimal bystander editing and negligible off-target activity—a critical advancement for therapeutic applications where safety and specificity are non-negotiable. This fidelity ensures that only the intended adenosine residues within target transcripts undergo conversion, thereby preserving global transcriptomic integrity while enabling precise modulation of gene function.

One illuminating application showcased in the study involves the endogenous CTNNB1 gene, which encodes β-catenin, a pivotal regulator in the Wnt signaling pathway. By employing PA-rABE to edit a phosphorylation site within CTNNB1 mRNA, researchers succeeded in stabilizing the β-catenin protein in vivo. Such stabilization potentiates the activation of canonical Wnt signaling, a pathway intricately involved in cell fate determination, proliferation, and tissue regeneration. This successful in vivo modulation testifies to the system’s functional versatility and capacity to induce physiologically relevant protein alterations through precise RNA editing.

The therapeutic potential of PA-rABE was further underscored through its application in a hemophilia B mouse model. Hemophilia B, a blood clotting disorder caused by mutations in the F9 gene coding for coagulation factor IX, was tackled by delivering an adeno-associated virus (AAV) vector encoding both PA-rABE and a variant of human F9 harboring a premature termination codon. Remarkably, upon blue light illumination, efficient RNA editing rescued the expression of functional clotting factor IX, leading to measurable amelioration of clotting defects in vivo. This represents a significant stride towards light-controlled gene therapies that could dramatically enhance patient safety and therapeutic outcomes.

The engineering of PA-rABE underscores a sophisticated interplay between molecular design and optical biology. By choosing to split the ADAR2 deaminase, the researchers cleverly rendered the enzyme inactive until illuminated, ensuring that baseline editing activity was undetectable in the absence of light stimulation. The Magnet dimerization system, adapted from engineered protein interaction domains, acted as a highly efficient optical switch, conferring rapid activation kinetics while allowing the system to return to an inactive state in darkness, thereby enabling reversible gene regulation.

Moreover, the choice of Cas13 as the RNA-targeting moiety brings additional advantages. Compared to widely used DNA-targeting CRISPR effectors, Cas13’s RNA recognition circumvents permanent genome alteration risks and can fine-tune protein expression levels without engendering inheritable genetic changes. Its compact size facilitates incorporation into current viral delivery vehicles, making it an ideal platform for therapeutic gene editing applications where controlled, transient intervention is desired.

In terms of delivery, the employment of adeno-associated virus vectors demonstrated practical feasibility for in vivo translation. AAVs remain the gold standard in gene therapy due to their low immunogenicity, ability to infect diverse tissues, and sustained transgene expression. Importantly, the modular design of PA-rABE enables packaging into a single or dual AAV construct, ensuring efficient co-delivery of both base editor components and target RNA transcripts. This scalability further enhances its translational promise.

The light-controlled dimension of PA-rABE opens exciting avenues for spatiotemporally defined gene regulation. Unlike small-molecule inducers, light can be precisely delivered to specific tissues or organ systems using fiber optics, endoscopy, or external illumination, enabling localized gene editing with minimal off-target systemic exposure. Furthermore, the rapid reversal of editor activation upon cessation of illumination provides dynamic modulation capabilities unattainable with irreversible genetic modifications.

From a safety perspective, the high target specificity and minimal off-target editing capacity reduce unintended transcriptomic perturbations, mitigating risks associated with RNA base editing. This meticulous control is vital for clinical applications, particularly when editing endogenous transcripts regulating critical cellular pathways such as Wnt signaling, whose dysregulation can promote oncogenesis if unchecked.

This breakthrough also carries profound implications for the broader field of RNA therapeutics. RNA base editors evade hurdles faced by DNA editing technologies, including double-strand break-induced genomic instability and permanent off-target mutations. PA-rABE’s reversible and controlled approach aligns seamlessly with growing interest in transient RNA modulation for treating genetic diseases, cancers, and neurological disorders.

The research team anticipates that further optimization of PA-rABE could refine its editing spectrum, enable multiplexed RNA targeting, and allow combination with other modalities such as RNA interference or translational modulation. Integration with advanced light delivery systems in clinical settings may establish PA-rABE as a versatile platform for precision gene therapy, regenerative medicine, and fundamental biology research.

In summary, the development of PA-rABE heralds a new era of photoactivatable RNA base editing, coupling the specificity and efficiency of A-to-I editing with unprecedented optical control. This technology elegantly addresses long-standing challenges in gene therapy by offering high-precision, reversible, and spatiotemporal modulation of gene expression. As this platform continues to evolve, it promises to unlock transformative biomedical applications and drive next-generation therapeutic strategies.

Subject of Research: Photoactivatable RNA base editing technology for precise, reversible, and spatiotemporally controlled gene modulation in vivo.

Article Title: Engineering a photoactivatable A-to-I RNA base editor for gene therapy in vivo.

Article References:
Li, H., Qiu, Y., Song, B. et al. Engineering a photoactivatable A-to-I RNA base editor for gene therapy in vivo. Nat Biotechnol (2025). https://doi.org/10.1038/s41587-025-02610-2

Image Credits: AI Generated

Tags: A-to-I RNA editorADAR2 deaminase technologyblue light stimulation in gene therapyCas13 RNA-targeting endonucleasecontrollable gene expressiongene therapy advancementsnovel gene editing technologiesphotoactivatable RNA editingphotodimerizable protein domainsreversible modulation of gene expressionRNA editing precisionspatiotemporal control in gene editing