‘Word processor’ for genes – Scientists uncover a fundamentally new mechanism for biological programming

Arc Institute scientists have discovered the bridge recombinase mechanism, a revolutionary tool that enables fully programmable DNA rearrangement.

Their find, described in detail in a recent Nature publication, is the first DNA recombinase to use non-coding RNA for sequence-specific selection of target and donor DNA molecules. This bridge RNA is programmable, allowing the user to specify any desired genomic target sequence and any donor DNA molecule to be inserted.

The research was developed in collaboration with the laboratories of Silvana Konermann, principal investigator of the Arc Institute and assistant professor of biochemistry at Stanford University, and Hiroshi Nishimasu, professor of structural biology at the University of Tokyo.

A new era of genetic programming

“The bridge RNA system is a fundamentally new mechanism for biological programming,” said Dr. Patrick Hsu, lead author of the study and principal investigator of the Arc Institute. University of California, Berkeley Assistant Professor of Bioengineering. “Bridge recombination can universally modify genetic material through sequence-specific insertion, excision, inversion, and more, enabling a word processor for the living genome beyond CRISPR.”

The bridging recombination system originates from insertion sequence 110 (IS110) elements, one of countless types of transposable elements—or “jumping genes”—that cut and insert to move within and between microbial genomes. Transposable elements are found in all life forms and have evolved into professional DNA manipulation machines to survive. IS110 elements are very minimal, consisting only of the gene encoding the recombinase enzyme and adjacent DNA segments that have remained a mystery until now.

The advanced mechanism of bridge RNA

Hsu’s lab found that when IS110 is excised from the genome, the non-coding ends of the DNA come together to form an RNA molecule—bridging RNA—that folds into two loops. One loop binds to the IS110 element itself, while the other loop binds to the target DNA where the element will be inserted. Bridge RNA is the first example of a bispecific guide molecule that specifies the sequence of both target and donor DNA through base-pairing interactions.

A team of researchers at the Arc Institute has discovered the bridge recombinase mechanism, a precise and powerful tool for recombining and rearranging DNA in a programmable manner. The bridging recombinase mechanism, which goes far beyond programmable genetic scissors such as CRISPR, allows scientists to specify not only the target DNA to be modified, but also the donor material to be recognized, so they can insert new functional genetic material, cut out defective DNA , or invert any two desired sequences. Discover more in this short video visualizing key aspects of the bridge recombination mechanism. Credit: Visual Science

Each loop of the bridge RNA is independently programmable, allowing researchers to mix and match any target and donor DNA sequences of interest. This means the system can go far beyond its natural role of inserting the IS110 element itself, instead allowing any desired genetic cargo—such as a functional copy of a faulty disease-causing gene—to be inserted into any genomic location. In this work, the team demonstrated more than 60% efficiency of insertion of the desired gene E-coli with more than 94% specificity for the correct location of the genome.

“These programmable bridge RNAs distinguish IS110 from other known recombinases, which lack an RNA component and cannot be programmed,” said co-author Nick Perry, a bioengineering graduate student at UC Berkeley. “It’s like the RNA bridge is a universal power adapter that makes the IS110 compatible with any outlet.”

Collaborative research and future implications

The Hsu laboratory’s discovery is complemented by their collaboration with the laboratory of Dr. Hiroshi Nishimasu at the University of Tokyo, also published on June 26 Nature. Nishimasu’s lab used cryo-electron microscopy to determine the molecular structures of the recombinase-bridged RNA complex bound to target and donor DNA, step-by-step through the key steps of the recombination process.

With further research and development, the bridging mechanism promises to usher in a third generation of RNA-directed systems that will expand beyond the CRISPR and RNA interference (RNAi) mechanisms for cutting DNA and RNA to offer a unified mechanism for programmable DNA rearrangement. Crucial to the further development of the bridging recombinase system for mammalian genome design, bridging recombinase joins both DNA strands without releasing cut DNA fragments, thereby overcoming a key limitation of current state-of-the-art genome editing technologies.

“The bridged recombination mechanism solves some of the most fundamental problems faced by other genome editing methods,” said lead researcher Matthew Durrant, principal scientist at Arc. “The ability to programmatically rearrange any two DNA molecules opens the door to breakthroughs in genome design.”

Reference:

“Bridge RNAs direct programmable recombination of target and donor DNA” by Matthew G. Durrant, Nicholas T. Perry, James J. Pai, Aditya R. Jangid, Januka S. Athukoralage, Masahiro Hiraizumi, John P. McSpedon, April Pawluk, Hiroshi Nishimasu , Silvana Konermann and Patrick D. Hsu, 26 Jun 2024, Nature.
DOI: 10.1038/s41586-024-07552-4

“Structural mechanism of bridging RNA-directed recombination” by Masahiro Hiraizumi, Nicholas T. Perry, Matthew G. Durrant, Teppei Soma, Naoto Nagahata, Sae Okazaki, Januka S. Athukoralage, Yukari Isayama, James J. Pai, April Pawluk, Silvana Konermann , Keitaro Yamashita, Patrick D. Hsu, and Hiroshi Nishimasu, 26 Jun 2024, Nature.
DOI: 10.1038/s41586-024-07570-2