Expanding the toolbox for RNA editing

Health problems are often caused by mutations that occur when genetic information is copied incorrectly, similar to a typo in a sentence. If these genetic typos are not corrected before they’re replicated, abnormal genetic changes can culminate in diseases like cancer.

In some cases, scientists can change DNA through a gene editing process using a tool called clustered regularly interspaced short palindromic repeats, known as CRISPR, which can be used to alter the genomes of living organisms.

RNA, the molecular cousin of DNA, serves as a messenger that carries copied excerpts of DNA to protein-making factories called ribosomes. Because RNA determines which proteins are expressed and ultimately how cells function, it is critical to understand processes involving RNA and find ways to correct erroneous RNA molecules. Among the variations of RNA, signals from guide RNA, or gRNA, attract the proteins that initiate editing activity.

Current RNA engineering techniques are limited to making only one type of edit at a time, limiting their usability. These techniques can have effects on areas not intentionally targeted, which can produce confounding results in scientific experiments or induce adverse side effects in treating disease.

Albert Cheng, an associate professor of biomedical engineering in the School of Biological and Health Systems Engineering, part of the Ira A. Fulton Schools of Engineering at Arizona State University, has developed a modular platform called Combinatorial RNA Engineering via Scaffold Tagged gRNA, or CREST, that modulates different RNA processing steps simultaneously — enabling several gene editing techniques to occur at a time — and reduces off-target events, the unintended changes at genomic sites that may lead to adverse outcomes. 

The CREST system has the potential to aid researchers in RNA-centric investigations and applications, like CRISPR. CREST can be used to adjust and edit RNA molecules in cells, thereby providing tools for scientists to study cellular function and potentially treat diseases.

In Cheng’s lab, researchers with extensive expertise in bioinformatics, next-generation sequencing and genetic engineering are focusing on the development of gene and RNA editing technologies to decipher the gene-regulating mechanisms underlying cellular reprogramming and disease stages.

A new tool for RNA editing 

Exploring RNA-centric methodologies is assisting in unraveling mysteries behind gene function as well as advancing opportunities for genetic engineering treatments to mitigate disease.

In a recent publication in the scientific journal Nucleic Acids Research, Cheng describes a new gene editing method to manipulate RNA processing steps. The technique allows researchers to control the production of molecules and treat malfunctions in RNA, which can prevent diseases.

He describes CREST as a way of advancing RNA from a single-task operating system to allow multiple procedures to run at the same time.

“We developed CREST to provide tools for precise tuning and editing of RNA molecules,” Cheng says. “Many diseases involve complex malfunctions of tens of different RNA molecules. The CREST platform allows multitasked operations to potentially correct many pieces of RNA at once.”

Cheng’s strategies for alternative splicing are expanding knowledge of RNA-binding proteins and enzymes. His team has identified proteins associated with guide RNA that travel throughout cells and demonstrated unprecedented multifunctional RNA modulation. 

“CREST can act like a socket with flexible functions and broad applications in RNA biology,” Cheng says.

Gene editing with intent

CREST transcends established methods by adding multiple docking points for RNA called scaffolds. The additional docking points provide more stations for editing, enabling several gene editing tools to be used simultaneously on different RNA molecules. 

The CREST platform also reduces off-target changes by splitting RNA editing enzymes into two parts and reassembling them on the guide RNA scaffold and the editing structure. The ability to reduce off-target effects highlights the significant advancements CREST provides as an RNA editing tool.

“In the future, we will create new functional modules for the platform so RNA can be tuned in different ways, creating an ‘app store’ for RNA biologists,” Cheng says.