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Phillip Zamore says that knocking out microRNAs in animals in the wild will instigate very different changes than in those raised in a controlled lab environment.
What lin-4 normally does is turn off another gene, lin-14, as Ambros realized after studying the interactions between lin-4 and lin-14 mutations. Gary Ruvkun, also a postdoc in Horvitz' lab, cloned lin-14 and found that it encoded a protein which was expressed in juvenile worms. Try as they might, however, Ambros and his colleagues could not find any protein that corresponded to the lin-4 gene.
Then came the breakthrough in 1993, recalls Horvitz. “In his own lab, Victor made the unexpected and remarkable discovery that lin-4 encodes a tiny, 21-nucleotide RNA. He and Gary further showed that this RNA—later recognized as the first microRNA—is complementary in sequence to its target, lin-14.” This observation, Horvitz says, led them to establish “a common mechanism of microRNA control of gene function—that microRNAs act by preventing the translation of their messenger target or targets.”
“The biomedical world, however, remained indifferent,” he notes. Most biologists viewed these molecules as just “weird little idiosyncratic things, limited to developmental timing in worms,” says Joan A. Steitz, an HHMI investigator at Yale University who studies other kinds of small RNAs and is now turning her eye to microRNAs. But seven years later, Ruvkun's lab cloned another mutant gene, let-7, which also encoded a 21-nucleotide RNA, and—most importantly—found that it is a very ancient gene, conserved in a wide range of animals from flies and sea urchins to humans. (It is the same let-7 microRNA that MIT's Jacks now hopes to use against lung cancer.) That discovery made scientists take notice.
Shortly afterward, in 2001, a blockbuster series of three articles was published in Science. In these articles, three separate labs headed by Ambros, Tuschl (then at the Max Planck Institute in Göttingen, Germany, and now an HHMI investigator at the Rockefeller University), and Bartel announced the existence of “a large class of tiny noncoding RNAs” with potentially broad regulatory functions in animals. This finding started a deluge of other newly identified microRNAs.
“We keep discovering more of them,” says Bartel, who estimates that humans have at least 500 different microRNAs, which regulate thousands of genes. “Back in 2001, we were happy when we had sequenced 300 small RNAs, 55 of which were unique microRNAs. Now, with high-throughput methods, we can sequence 5 million small RNAs at a time, from humans or any animal we choose. This gives us the ability to find microRNAs that we'd missed earlier.” Those they found early on are highly conserved through evolution, according to Bartel. “As we dig deeper, we get those that are expressed at lower levels and are less likely to be conserved in other animals,” he says.
Nevertheless, in a paper published October 30, 2008, in Nature, Bartel and colleagues at MIT; the University of California, Berkeley; and the University of Queensland showed the influence of microRNAs throughout history. “MicroRNAs have been available to regulate and shape gene expression as far back as we can go in animal evolution—they might even predate animals,” he says. “They might have helped to usher in the era of multicellular animal life.”
Photo: Robert E. Klein / AP, ©HHMI