Shut up, Gen, shut up!
Scientists have improved the gene suppression tool
Nanonewsnet based on IMP materials: A Molecular Toolkit for Gene Silencing
The group of Johannes Zuber, MD, from the Institute of Molecular Pathology (IMP), Austria, managed to eliminate key limitations in the use of RNA interference (RNAi), a unique method of suppressing gene expression.
By improving the structure consisting of hairpin RNA and microRNA, scientists were able to significantly increase the efficiency and accuracy of gene silencing (from the English silence – to be silent). In an optimized form, the method facilitates the search for drug targets and allows for more accurate interpretation of experimental results. The Institute of Molecular Pathology intends to make this RNAi toolkit available to all interested scientists. The results of the work of the Austrian researchers are published in the latest issue of the journal Cell Reports: An optimized microRNA backbone for effective single-copy RNAi.
RNA interference (RNAi) is a regulatory mechanism naturally present in biological cells. Short fragments of RNA (the so-called "hairpins", shRNAs) come into conflict with the genetic information read from DNA and recorded in the matrix RNA, preventing protein synthesis and, thus, silencing genes. In 1990, this phenomenon was first discovered in plants, and in 2001 – in mammals. Scientists immediately realized the enormous potential of this mechanism as an effective means of experimental suppression of gene activity. Apart from numerous applications in fundamental biological research, RNA interference has become a unique method for identifying and studying therapeutic target genes. Nevertheless, despite the enormous possibilities, the currently available RNAi reagents often turn out to be ineffective or have non-specific side effects.
The idea of how to improve RNAi technology came to Dr. Zuber and his colleague Christof Fellmann back in 2010, when they both worked at Cold Spring Harbor Laboratory in the USA.
"The basic principles of RNA interference have not yet been fully studied. To turn off a certain gene, you need to test a lot of hairpin molecules, and often only one out of ten is effective enough. To improve the method, we took nature itself as an example," explains Dr. Zuber's train of thought.
A particularly powerful and widely used method of RNA interference is based on the embedding of synthetic hairpin sequences into the natural skeletons of microRNA molecules. As a result, RNA constructs are formed that mimic natural structures that are processed by normal cellular pathways. However, the effectiveness of the reagents obtained in this way leaves much to be desired.
Dr. Zuber and his team analyzed the skeletons of human microRNAs, focusing on those parts of the sequences that have not changed during evolution – a sign that they can perform important functions. The scientists realized that in the synthetic microRNA skeletons commonly used in the creation of shRNAmiRs constructs, some of these sequences were altered. By eliminating these differences and systematically testing many variants, Zuber and his colleagues significantly improved the effectiveness of the synthetic RNA interference tool.
"The benefits for science are huge," Dr. Zuber emphasizes the significance of his results. While up to twenty pins need to be tested for powerful suppression of this gene by existing methods, optimized reagents reduce their number to an average of four.
Diagram from an article in Cell Reports – VM
In the future, Zuber's research will allow better use of the potential of RNA interference in cancer research. Despite huge efforts, pharmaceutical companies have not yet managed to develop a single drug based on RNAi. Nevertheless, large-scale RNAi screenings represent a unique method of finding and testing the most promising target genes for new drugs before launching the slow and expensive process of developing a new drug. The new optimized RNAi reagents will be especially useful in such highly productive screenings, as they will allow more genes to be tested simultaneously and will do so with increased efficiency and accuracy. And, most importantly, the introduction of optimized RNAi tools will reduce the risk of missing promising targets for future therapeutic use.
Portal "Eternal youth" http://vechnayamolodost.ru23.12.2013