Scientists have known for more than 20 years that all cells use a strategy called RNA interference to regulate gene expression, but the new study describes for the first time how specific proteins manage the step-by-step process of assembling molecular complexes that play regulatory roles.
Among the surprising discoveries is the suppression of the interference’s target messenger RNA (mRNA), a protein whose instructions are carried by this same mRNA, which also helps complete the final step in assembling the complex that inhibits gene expression.
Overall, this study shows that management proteins transform precursor complexes into mature RNA-induced silencing complexes (RISCs) through a four-step process.
“This mechanism has been a black box for a quarter of a century since RNA interference was discovered,” said lead author Kotaro Nakanishi, a professor of chemistry and biochemistry at The Ohio State University.
“Visualizing mechanisms is very important. Pharmaceutical companies may appreciate our 3D structures because they can optimize and design new drugs based on them.”
The study was published today (May 26, 2026) in the journal. molecular cell.
In a 2022 paper, Nakanishi outlined how a “superfamily” of four Argonaute proteins is involved in RNA interference, explaining that microRNAs and small interfering RNAs (siRNAs) are both small segments of RNA that inhibit the protein-building functions of genes, and must be loaded into Argonaute proteins to perform their job; they cannot do their job alone.
In this new study, Nakanishi and colleagues performed biochemical assays and cryo-electron microscopy using Argonaute2 as a model. “But based on biochemical data, we believe that all Argonaute proteins behave in the same way,” he said.
The starting point for the study involved incubating specific siRNA duplexes with human Argonaute2 protein.
As a result, a series of subsequent steps were identified. Argonaute2 loads double-stranded RNA and selects one strand over another to act as a guide. The protein then unwinds the duplex and expels the “passenger” strand, which is no longer needed.
This evacuation revealed a surprising role for the target mRNA (helping to shed the passenger strand), leading the team to call the final step TAPE (target-assisted passenger ejection).
Although in most cases of RNA interference, mRNA has been considered primarily as a target for microRNAs and siRNAs, “our results instead suggest that mRNA may also bind to precursor RISC and facilitate passenger removal during RISC maturation,” the authors write.
This is complicated, but it also makes sense to me that if cells are targeting key mRNAs, these target mRNAs will also promote RISC formation. ”
Kotaro Nakanishi, Professor of Chemistry and Biochemistry, Ohio State University
He said the findings should help advance the development of therapeutic siRNAs and cityRNAs (cleavage-induced small RNAs) that can be engineered to trigger specific RNA interference processes that override natural cellular processes and silence problematic genes associated with diseases.
“Researchers and pharmaceutical companies have used siRNA as a potential therapy to block gene expression and study the role of proteins of interest. However, no one knows how RISCs are formed,” Nakanishi said. “Now we can provide a robust structural foundation or foundation on which to design or optimize siRNAs.”
His lab is now using the same technique to see how three other Argonaute proteins assemble RISC.
This research was supported by fellowships from the National Institutes of Health and the Center for RNA Biology at Ohio State.
Co-authors include Huaqun Zhang, Vishal Annasaheb Adhav, Audrey Kehling, Andrew Savidge, and Zhangfei Shen, all of Ohio. Giovanna Grandinetti and Yohie Narui of the Ohio Electron Microscopy Analysis Center helped collect high-quality cryo-EM micrographs.
Mr. Nakanishi is a co-founder and scientific advisor of City Therapeutics, Inc. and reports a financial interest in it.
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Reference magazines:
Chan, H. others. (2026). Structural basis of human Argonaute2 RISC assembly. molecular cell. DOI: 10.1016/j.molcel.2026.04.029. https://www.sciencedirect.com/science/article/pii/S1097276526002832?via%3Dihub
