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The new technique uncovers details about protein production that had been technologically out of reach. To grasp how cells function or run awry, scientists would ideally like to directly monitor their proteins, but that’s technically tough to do. The best proxy has been to measure amounts of different protein-coding mRNAs in the cell by using existing microarray tools; those techniques, however, can’t identify which mRNA messages actually get converted into proteins. Because ribosome profiling goes a step further and quantifies that missing information with a massive amount of data, it represents “a really major advance that will become widely used,” says geneticist and ribosome expert John F. Atkins of University College Cork in Ireland.
The Nature study of ribosome pausing is “a nice example” of the technique’s power, Atkins says. Weissman’s team scanned the mRNAs in bacterial cells, looking for all the places where ribosome footprints pile up, like in a traffic jam. Their computational analysis found “a lot more pauses than people expected,” Li says—occurring at hundreds to thousands of short mRNA sequences.
Those short stretches all shared a similarity to the so-called Shine-Dalgarno motif, found in many bacteria. It’s a special sequence that the ribosome recognizes and binds to before it starts reading a protein-coding message just downstream. But the message itself can also include consecutive codons that together form a sequence that is Shine-Dalgarno-like and which had been known in a few specialized cases to cause pausing. Weissman’s team demonstrated that these Shine-Dalgarno look-alikes are the global mechanism behind pauses. “They were sort of sticking like Velcro to the ribosome and slowing things down,” Weissman says.
The pausing mechanism helps to explain why nature evolved genomes to have different preferences for different codon synonyms: they confer various advantages or disadvantages for protein production. For example, the Weissman team’s analysis found that bacterial genomes contain relatively few of the pause-inducing codons, Li says. That is because the bugs evolved to grow fast, which requires rapidly churning out proteins. However, pauses are sometimes desirable for key purposes, such as to regulate protein synthesis and, possibly, to let an elongating protein undergo partial folding, the researchers say.
Weissman began investigating protein production only five years ago, when he decided to develop ribosome profiling in part to tackle a question about protein-folding, then his main career focus. His father, Sherman, a prominent Yale University geneticist, was a great source of advice during the shift, both on big-picture strategizing and on the nuts and bolts of using superfast next-generation DNA sequencers.
As Weissman read up on the intricacies of ribosomes, another exceptional sounding board was Marshall Nirenberg, who late in life married Weissman’s mother, Myrna, in 2005. That’s how Weissman ended up with one of the six existing copies of Nirenberg’s genetic code notes. (The originals are at the National Library of Medicine.) Although the Nobel laureate had long ago switched to neurobiology research, he told Weissman that he had been grappling with questions about codon synonyms and ribosome reading speeds from the very beginning.
“That gave me a context and a sense of the importance of the problem,” Weissman says—and motivated him to apply his profiling technique to finding answers. The method could have a far broader impact on molecular biology research than just for protein folding studies, he realized.
Indeed, from both his superstar advisors, Weissman says he learned a pivotal lesson: new scientific tools are valuable not just for making research faster or easier but also for potentially opening the door to fundamental, game-changing discoveries. The genetic code notes on his wall remind him of that every day. They are “an inspiration to work on problems that are broad and basic and that can touch a lot of different fields,” he says.