Arc Institute scientists have discovered the bridge recombinase mechanism, a revolutionary tool that allows complete programming of DNA rearrangements.
Their discoveries are detailed in a recent publication Natureis the first DNA recombinase to use noncoding 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 added.
The work was developed in collaboration with the laboratories of Silvana Konermann, principal investigator at the Arc Institute and associate professor of biochemistry at Stanford University, and Hiroshi Nishimasu, professor of structural biology at the University of Tokyo.
A new era in genetic programming
“The bridging RNA system is a fundamentally new mechanism for biological programming,” said Dr. Patrick Hsu, senior author of the study and principal investigator of the Arc Institute and associate professor of bioengineering at UC Berkeley. “Bridging recombination can universally modify genetic material through insertions, deletions, inversions, etc., creating a word processor for the living genome beyond CRISPR.”
The bridging recombination system is derived from insertion sequence 110 (IS110) elements, one of numerous types of transposable elements (or “jumping genes”) that cut and insert themselves to move within and between microbial genomes. Transposable elements are found in all life forms and have evolved into professional DNA manipulation machines for survival. IS110 elements are quite minimal and consist only of the gene encoding the recombinase enzyme and flanking DNA segments that have remained mysterious until now.
Enhanced bridging RNA mechanism
Hsu’s lab found that when IS110 is removed from the genome, the noncoding ends of the DNA come together to form an RNA molecule that folds into two loops (a bridge RNA). One loop binds to the IS110 element itself, while the other loop binds to the target DNA to which the element will be added. Jumper RNA is the first example of a bispecific guide molecule that specifies the sequence of both target and donor DNA through base-pairing interactions.
Each loop of bridge RNA can be programmed independently, allowing researchers to mix and match any target and donor DNA sequence of interest. This means the system can go far beyond its natural role of inserting the IS110 element, 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 study, the team demonstrated over 60% efficiency in introducing the desired gene into the body. Escherichia coli It has over 94% specificity for the correct location in the genome.
“These programmable bridge RNAs distinguish IS110 from other known recombinases that lack an RNA component and cannot be programmed,” said lead author Nick Perry, a graduate student in bioengineering at the University of California, Berkeley. “It’s as if the bridge RNA were a universal power adapter that makes IS110 compatible with any outlet.”
Collaborative research and future implications
Hsu’s lab’s discovery complements their collaboration with Dr. Hiroshi Nishimasu’s lab at the University of Tokyo, also published June 26. NatureNishimasu’s lab used cryo-electron microscopy to determine the molecular structures of the recombinase-bridge RNA complex that binds to target and donor DNA and progresses sequentially through key steps in the recombination process.
With further research and development, the bridge mechanism promises to usher in third-generation RNA-guided systems, moving beyond the DNA and RNA cutting mechanisms of CRISPR and RNA interference (RNAi) to offer a unified mechanism for programmed DNA rearrangement. Critical to further developing the bridge recombination system for mammalian genome design, bridge recombinase joins both DNA strands without releasing the cut DNA fragments, bypassing a major limitation of current state-of-the-art genome editing technologies.
“The bridging recombination mechanism solves some of the most fundamental problems faced by other genome editing techniques,” said Matthew Durrant, a senior scientist at Arc and one of the leaders of the study. “The ability to programmatically rearrange any two DNA molecules opens the door to a breakthrough in genome design.”
Source: Port Altele
