In two papers published in the latest Science, scientists from Howard Hughes Medical Institute describe a way of identifying genes that are critical to colon and breast cancer cell growth. The technique should help with developing new therapeutic methods to target these common cancers.
The method exploits a powerful cellular mechanism called RNA interference. Discovered just a decade ago, RNA interference likely evolved to help cells fight viruses. The cellular machinery involved in RNA interference first identifies short segments of suspicious-looking RNA, and then destroys all identical copies of that RNA. The result: None of the protein that the RNA encodes for gets made.
While RNA interference prevents viruses from replicating inside cells, scientists discovered that they could exploit the process to squelch individual gene products. To do so, they introduce a short segment of RNA that looks like one of the cell’s normal genes. The RNA interference machinery grinds into action and shuts down production of the protein made from that gene.
Six years ago, Elledge and Hannon began making a library of RNAs, called short hairpin RNAs, which trigger the RNA interference mechanism. They’ve now made short hairpin RNAs that can squelch every gene in the human and mouse genomes.
For their new experiments, the pair first identified about 3,000 genes important in cell signaling, growth, and other essential processes. Next, they inserted a gene sequence coding for short hairpin RNAs targeting these genes into retroviruses. Then they infected dishes of normal and tumor cells with the retroviruses, which added instructions to each cell’s genome telling it to produce a short hairpin RNA. These short RNAs then triggered the RNA interference mechanism. In effect, each virus halts production of a single protein in a single cell.
In the past, researchers deployed this method to study the effects of turning off one particular gene. But to study the effects of thousands of genes, researchers had to run thousands of separate experiments with thousands of plates of cells.
Instead, Hannon and Elledge developed a “barcoding” method to track a diverse pool of short hairpin RNAs in parallel. In the barcoding method, every short hairpin RNA that is made carries a unique genetic tag. This tag lets the researchers track the effect of thousands of the RNAs in a single pool of cells in a single lab dish.
“We get a mixture of cells where each individual cell has one of these genes knocked down,” says Hannon.
If RNA interference knocks down a gene important for cell growth and survival, the cell fails to thrive or dies. At the end of the experiment, the researchers recover only small amounts of the short hairpin RNA associated with that gene. The researchers then know that the gene is a potential Achilles heel for the cell.
In the research reported in the Science papers, the scientists ran many such experiments on normal, breast, and colon cancer cells. The team found dozens of genes that, when eliminated, hinder cancer cells but don’t seem to harm normal cells.
Watch a short clip explaining the technology.
Press release: New Genetic Barcoding Technique Identifies Dozens of Targets for Cancer Drugs
Abstracts in Science Magazine: Profiling Essential Genes in Human Mammary Cells by Multiplex RNAi Screening; Cancer Proliferation Gene Discovery Through Functional Genomics