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In RNA interference, double-stranded ribonucleic acid (dsRNA) targets destruction of messenger RNA (mRNA) that has the same sequence it has. The Dicer enzyme cuts dsRNA into small inhibitory RNA (siRNA), which lures mRNA into the RISC (RNA-induced silencing complex) protein complex where it is destroyed.
Mello became fascinated with one science after another as he was growing up—paleontology, evolution, geology, astronomy—but he traces his interest in molecular biology to June 12, 1978. A few days before graduating from high school, he read an article in the Washington Post titled "Bacterium Is Used in Producing Insulin." It described one of the first applications of genetic engineering: the insertion of a human gene into bacteria to produce insulin for diabetics. "I was absolutely amazed to learn that bacteria could read the human genetic code," Mello says. "I realized that I would love to be able to understand how genes function by taking them out and putting them back into an organism."
Like their parents, Mello and his older brother and sister had gone to Brown University. (Roger, the only exception in the family, attended the University of Virginia on a wrestling scholarship after becoming state champion in high school.) After Brown, Mello went to graduate school at the University of Colorado at Boulder, where he began the research that would lead to his Nobel Prize. Working in the laboratory of David Hirsh, he focused on techniques to introduce genes into a widely used model organism, the tiny worm Caenorhabditis elegans. Around that time, Andrew Fire, who was doing graduate and postdoctoral work at the Massachusetts Institute of Technology and the Medical Research Council Laboratory of Molecular Biology in England, developed a way to microinject DNA directly into the nucleus of a worm egg. But the technique was so difficult that Mello developed an alternative microinjection strategy, and in the process the two got to know each other and began collaborating. "He was very gracious," says Mello. "The worm community is a very sharing community."
After more graduate work at Harvard University and a postdoctoral fellowship at Seattle's Fred Hutchinson Cancer Research Center, Mello set up his own laboratory at the University of Massachusetts Medical School in 1994. The same year, a graduate student at Cornell University named Su Guo made a puzzling discovery. She was injecting molecules known as antisense RNA into C. elegans to block the action of a particular gene. (Antisense RNA molecules are designed to bind with single-stranded RNA molecules that have an exactly opposite genetic sequence.) But when Guo injected "sense" (or same-sequence) RNA—which should not bind with messenger RNA—into the cell to establish a control, the action of the gene also was blocked. "That was a surprise," says Mello. "But everyone assumed that there was antisense contaminating the sense RNA preparation."
Mello's lab worked primarily on the developmental biology of C. elegans, but he was so intrigued by Guo's observation that he launched a separate project to explore it. One day, after injecting antisense RNA into C. elegans, he got distracted and did not check the worms until after they had reproduced. Amazingly, he found that their progeny also exhibited gene silencing, even though there was no obvious way for the effect to travel from one generation to the next. "That to me was stunning, unbelievable," he says.
Not long afterward, another fortunate accident occurred. Sam Driver, a new graduate student in Mello's lab, was trying to inject RNA into particular cells and kept missing. Yet even when he hit cells that weren't anywhere near his target, the gene silencing spread from one cell to another, though there was not nearly enough RNA for the effect to be so widespread. "We had no clue what was going on," Mello recalls.
Fire, who was then at the Carnegie Institution of Washington's Department of Embryology in Baltimore, had been conducting similar experiments, and he and Mello often compared results over the phone and at meetings. One day Fire made an off-the-wall suggestion. Maybe the preparations of single-stranded RNA also contained small amounts of double-stranded RNA, and the double-stranded RNA was causing the effect. The idea seemed improbable. Double-stranded RNA should be relatively inert in a cell, whereas single-stranded RNA would bind to complementary strands. But Mello was beginning to wonder if the single-stranded RNA was working alone in C. elegans. The only way for the effect to spread from cell to cell or from parent to offspring was if it was being amplified by some sort of cellular machinery.
Fire's idea cracked open the case. When he isolated double-stranded RNA from the preparations of single-stranded DNA, he found that the double-stranded RNA had a much stronger silencing effect than either the antisense or the sense single-stranded RNA.