ABA-Induced Cargo Proteins Loading in Extracellular Vesicles for Gene Editing

TL;DR

Researchers developed a system using abscisic acid to control the loading of specific proteins, like Cas9, into exosomes for targeted delivery and gene editing.

Contribution

The study introduces an ABA-inducible system for programmable and controllable loading of therapeutic proteins into exosomes.

Findings

The BASP1–PYL1 fusion was the most effective scaffold for ABA-dependent protein loading into exosomes.

ABA administration successfully triggered the encapsulation of functional cargo proteins, including Cas9, into extracellular vesicles.

Loaded exosomes retained their canonical characteristics and delivered functional cargo to recipient cells, enabling genome editing.

Abstract

What are the main findings?
We developed an ABA-inducible proximity system that successfully directs theencapsulation of specific protein cargo (e.g., Cas9) into exosomes during biogenesis.Among the engineered scaffolds, the BASP1–PYL1 fusion proved most effective, enabling robust, ABA-dependent protein loading into EVs.

We developed an ABA-inducible proximity system that successfully directs theencapsulation of specific protein cargo (e.g., Cas9) into exosomes during biogenesis.

Among the engineered scaffolds, the BASP1–PYL1 fusion proved most effective, enabling robust, ABA-dependent protein loading into EVs.

What are the implications of the main finding?
This work establishes a versatile molecular switch for controllable loading of therapeutic proteins into exosomes.The technology provides a programmable platform for next-generation therapeutic delivery.

This work establishes a versatile molecular switch for controllable loading of therapeutic proteins into exosomes.

The technology provides a programmable platform for next-generation therapeutic delivery.

Extracellular vesicles, which carry bioactive cargos such as proteins, RNAs, and lipids, represent promising drug delivery vehicles owing to their biocompatibility, low immunogenicity, and inherent tissue-targeting capabilities. To address the current limitations in controlled cargo loading, we developed an abscisic acid (ABA)-inducible proximity system that directs proteins into exosomes during biogenesis. We engineered exosomal scaffolds by fusing the ABA receptor PYL1 to EV-enriched proteins—including BASP1, CD9, PTGFRN, and a truncated form PTGFRNΔ687—thereby creating docking sites within the exosomal lumen, while the target cargo (e.g., EGFP, firefly luciferase, or Cas9) was tagged with the ABI1 phosphatase domain. We demonstrate that ABA administration in producer cells induces PYL1–ABI1 complex formation, which recruits ABI1-fused cargo for selective encapsulation into EVs. Among the scaffolds tested, BASP1–PYL1 proved the most effective, enabling robust, ABA-dependent enrichment of cargo proteins. Purified EVs maintained canonical morphology, size, and marker expression (CD63, syntenin-1, CD9), confirming preserved biogenesis. Critically, these loaded exosomes efficiently delivered functional cargo to recipient cells, enabling Cas9/sgRNA-mediated genome editing. Together, our findings establish an ABA-triggered molecular switch for controllable EV protein loading, providing a versatile platform for next-generation therapeutic delivery.

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