More findings confirm that small RNAs work in mysterious ways.
Nearly 20 years after its discovery, RNA interference (RNAi) is part of biology’s orthodoxy. Small RNA molecules can disrupt gene expression by degrading messenger RNAs (mRNAs) on their way to becoming proteins, or otherwise interfering with translation. But the discovery that these same small RNA molecules might be able to do just the opposite—enhance gene expression—was somewhat heretical.
In 2007, molecular biologist Shobha Vasudevan of Yale University and her colleagues produced the unanticipated findings: Small RNA molecules known to be involved in RNAi, known as microRNAs (miRNAs), can activate translation, promoting the conversion of mRNAs to proteins. It was a “surprise finding,” Vasudevan recalls.
Further investigation revealed that activation occurred only during cell-cycle arrest, induced by serum starvation. In actively growing cells, on the other hand, miRNAs suppressed translation. The exact mechanism of activation is unclear, Vasudevan says, but it appears to involve the recruitment of Argonaute (AGO) proteins—known participants in the RNAi pathway—and fragile X mental retardation–related protein 1 (FXR1). These proteins combine with the miRNA to form complexes that bind to the 3' untranslated region (3'-UTR) of the mRNA of tumor necrosis factor-α (TNFα) to initiate translation.
This discovery was hot on the heels of another unexpected finding—that of RNA activation (RNAa) at the level of gene transcription. Just one year earlier, molecular biologists Long-Cheng Li and Robert Place and their colleagues at the University of California, San Francisco, found that small RNAs could switch on gene transcription1—a finding that was corroborated just a few months later by a group of researchers working independently at the University of Texas Southwestern Medical Center at Dallas.2 (See the May 2009 issue of The Scientist.) And last year, additional research revealed more of the mysterious qualities behind translation activation by miRNA.
“These papers are making us look at miRNAs as a far more versatile system of regulating gene expression,” says Vasudevan, now at Massachusetts General Hospital and Harvard Medical School.
Five months after the publication of this month’s hot paper, Anders Lund of the University of Copenhagen in Denmark and his colleagues discovered another example of miRNA-mediated translation activation—this time of mRNAs that encode ribosomal protein.3 “We were looking for just general targets for miR-10a”—an miRNA broadly expressed in mice, Lund recalls. “What we found, which was very much a surprise,” was that miR-10a interacts with several ribosomal protein mRNAs to stimulate their translation. Even more bizarre, however, was the target: miR-10a appeared to be acting on the 5'UTR of ribosomal mRNA—the so-called 5'-terminal oligopyrimidine tracts, or 5'TOP motif. miRNAs predominately suppress gene expression by binding to target sites in the 3'UTR of mRNAs—the end of the mRNA strand where translation is completed.
It doesn’t make sense, Lund says. Placing an miRNA complex in between the 5' mRNA cap—which normally recruits the small ribosome to initiate translation—and the AUG start codon seems “obstructive,” Lund says, “unless [there is] some completely different mechanism.”
Later in 2008, yet another example of miRNA translation activation emerged. In a study led by virologist Michael Niepmann of Justus-Liebig-University Giessen in Germany, the researchers discovered that the liver-specific miR-122 could stimulate translation of hepatitis C virus (HCV) mRNAs. In this case, miR-122 interacted with two target sites to enhance the association of the viral RNA with the host ribosomes.4 The targets were, again, at the 5' end of the mRNA. Most recently, this October, researchers published a third report of 5'UTR translation activation, in which miR-346 targets the 5'UTR of receptor-interacting protein 140 (RIP140), a transcriptional corepressor.5
Whether targeting the 5'UTR is a common mechanism of miRNA translation activation, “it’s way too early to say,” Lund says. Indeed, this past summer, biochemist Yukihide Tomari of the University of Tokyo and his colleagues showed that miRNAs targeting the 3'UTR activated translation under certain conditions—specifically, when the target mRNA lacks a poly(A) tail, present at the 3' end. “We are (almost hopelessly) puzzled as to mechanistically how such translational activation can occur,” Tomari says in an email.
“I think it is clear that there’s no universal mechanism,” says RNAi researcher Timothy Nilsen of Case Western Reserve University in Ohio, “and it’s going to be important to find out in what context you can either up-regulate or down-regulate [gene expression].”
The discovery that miRNAs can enhance gene expression has captured the attention of the biotechnology sector. Place, for one, has sold his technology regarding miRNA activation of transcription to Massachusetts-based Alnylam Pharmaceuticals for therapeutic development, and is currently looking to license the technology for reagent development to a company that he declined to name.
In September of this year, molecular biologist Seppo Ylä-Herttuala of the University of Kuopio in Finland and Ark Therapeutics, along with his colleagues, published the first demonstration that small RNAs can be delivered by a lentriviral vector to up-regulate transcription in vivo.6 “I’m pretty sure that this is a general mechanism that is used for fine-tuning gene expression at the nuclear level,” Ylä-Herttuala says, meaning that it could have broad applications for developing reagents as well as (eventually) therapeutic treatments.
Place sees just as much potential for the up-regulation of translation, which evades epigenetics by targeting mRNAs in the cytoplasm. “We don’t know [yet] to what extent we can exploit translation activation,” Place says, “[but] if it can be [broadly applied] to any given transcript, that technology is essentially limitless.”
|Data derived from the Science Watch/Hot Papers database and the Web of Science (Thomson ISI) show that Hot Papers are cited 50 to 100 times more often than the average paper of the same type and age.
S. Vasudevan, et al., “Switching from Repression to Activation: MicroRNAs Can Up-regulate Translation,” Science, 318:1931–34, 2007. (Cited in 250 papers)