MADRID, June 28 (EUROPA PRESS) –
In a leap forward for genetic engineering, a team of researchers at the Arc Institute (United States) has discovered the bridging recombinase mechanism, a precise and powerful tool to recombine and rearrange DNA in a programmable way.
The study, published in Nature by researchers at the University of Berkeley (United States), reports the discovery of the first DNA recombinase that uses a non-coding RNA for sequence-specific selection of target and donor DNA molecules. This RNA bridge is programmable, allowing the user to specify any desired genomic target sequence and any donor DNA molecule to be inserted.
“The RNA bridging system is a fundamentally new mechanism for biological programming,” reports Hsu, lead author of the study and principal investigator at the Arc Institute and assistant professor of bioengineering at UC Berkeley in the United States. “Bridging recombination can universally modify genetic material through sequence-specific insertion, excision, inversion, and more, enabling a word processor for the living genome beyond CRISPR.”
Arc senior scientist Matthew Durrant and UC Berkeley bioengineering graduate student Nick Perry were lead authors of the discovery. The research was developed in collaboration with the laboratories of Silvana Konermann, principal investigator at the Arc Institute and assistant professor of biochemistry at Stanford University (United States), and Hiroshi Nishimasu, professor of structural biology at the University of Tokyo (Japan).
The bridging recombination system comes from insertion sequence 110 (IS110) elements, one of countless types of transposable elements – or jumping genes – that cut and paste to move within and between microbial genomes. Transposable elements are found in all forms of life and have evolved into professional DNA manipulation machines for survival. The IS110 elements are minimal, consisting only of a gene encoding the recombinase enzyme, plus flanking DNA segments that, until now, remain a mystery.
Hsu’s lab discovered that when IS110 is cleaved from a genome, the ends of the noncoding DNA join together to produce an RNA molecule (the 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. The bridging RNA is the first example of a bispecific guide molecule, which specifies the sequence of the target and donor DNA through base-pairing interactions.
Each loop of the bridging RNA is independently programmable, allowing researchers to mix and match any DNA sequence of interest with the target and donor. This means that the system can go far beyond its natural function of inserting the IS110 element, instead allowing the insertion of any desirable genetic cargo, such as a functional copy of a defective disease-causing gene, at any genomic location. In this work, the team demonstrated more than 60% insertion efficiency of a desired gene into E. coli with more than 94% specificity for the correct genomic location.
“These programmable bridging RNAs distinguish IS110 from other known recombinases, which lack an RNA component and cannot be programmed,” notes graduate student Nick Perry. “It’s as if the RNA bridge is a universal power adapter that makes the IS110 compatible with any outlet”
The Hsu lab’s discovery is complemented by their collaboration with Dr. Hiroshi Nishimasu’s lab at the University of Tokyo, also published in Nature. Nishimasu’s lab used cryo-electron microscopy to determine the molecular structures of the recombinase-bridge RNA complex bound to target and donor DNA, progressing sequentially through key steps of the recombination process.
With further exploration and development, the bridging mechanism promises to usher in a third generation of RNA-guided systems, expanding beyond the DNA and RNA cleavage mechanisms of CRISPR and RNA interference (RNAi) to offer a unified mechanism for programmable DNA rearrangements. Critical to the further development of the bridging recombination system for mammalian genome design, the bridging recombinase joins both DNA strands without releasing cut DNA fragments, circumventing a key limitation of state-of-the-art genome editing technologies.
“The bridging recombination mechanism solves some of the most fundamental challenges faced by other genome editing methods,” concludes research co-leader Durrant. “The ability to programmably rearrange two DNA molecules opens the door to advances in genome design.”
