What is the best way to discover how a gene works when it is on? In many cases, the answer is to turn it off.
Gene silencing is the process by which gene expression, or the proper “effect” stimulated by the activation of a gene, is diminished or prevented completely. In plants and animals, this is a naturally occurring process that serves important roles, including shielding organisms from harmful invaders such as viral agents. In the past three decades, scientists have developed several methods of artificially harnessing this natural path to be used as a tool in medical research, gene therapy, and agriculture.
A key discovery in gene silencing was made in 1998 when Andrew Fire first identified RNA interference (RNAi) — a process of silencing genes post–transcription. Small interfering RNAs (siRNAs) complementary to the target gene’s transcript sequence are incorporated into an RNA–induced silencing complex. This complex degrades a messenger RNA (mRNA) target, halting gene expression. This revelation led to the first systems for targeted gene silencing, and, as of May 2024, RNAi is actively used in plants for antiviral defense, pest control, and improving drought resistance.
A method of gene silencing introduced in 2013 harnesses the power of clustered regularly interspaced short palindromic repeats, more commonly referred to by their acronym, CRISPR. CRISPR and associated enzymes form the foundation of a system that precisely modifies genetic information through cleaving DNA at specific locations. When a gene is targeted at its promoter sequence, RNA polymerase and transcription factors cannot bind; by extension, gene expression is reduced or diminished completely.
“A method of gene silencing introduced in 2013 harnesses the power of clustered regularly interspaced short palindromic repeats, more commonly referred to by their acronym, CRISPR.”
In a 2015 study, researchers used CRISPR interference to target genes in mycobacterium in order to understand Mycobacterium tuberculosis (MtB), the bacterium responsible for the highly infectious tuberculosis disease. This is just one of many examples of CRISPR technology in action. Researchers implemented a CRISPRi system in the fast-growing model M. smegmatis (Msm), and the slow–growing MtB bacteria, using codon–optimized dCas9 of S. pyogenes and a constructed E. coli–mycobacteria shuttle plasmid to efficiently repress multiple genes simultaneously and test gene essentiality in mycobacteria. In this study, researchers were able to identify the gene EngA as essential for the growth of Msm, creating an avenue for a potential tuberculosis treatment in the future.
In December 2023, the first FDA approved CRISPR cell–based gene therapy, Casgevy, was approved as a treatment for sickle cell disease. In sickle cell disease, a mutation in hemoglobin causes a disfigurement of red blood cells, leading to reduced oxygen delivery, severe pain and organ damage in the form of vaso–occlusive crises. In order to treat this damage, scientists modify blood stem cells in patients, and then transplant them back into the patient where they attach and multiply, increasing the number of hemoglobin proteins facilitating oxygen delivery. This technology is already being utilized, and according to the New York Times, the very first adolescent to receive this therapy was reported to have finished treatment in mid–October 2024.
Gene silencing may seem like a quiet mechanism, but its impact is loud. As technologies continue to develop, gene silencing applications are becoming more widespread, revealing more about genomes and paving the way for future medical breakthroughs.
