Meet a dicey enzyme named Dicer (pun intended), which chops away at sequences of RNA - ribonucleic acid. Dicer performs its slicing gig in every major species of life on earth. It operates on insects, plants, worms, birds and mammals - including humans.
"Dicer is one of the enzymes involved in processing small interacting RNAs," explained Jean-Marc Jacque, an immunologist at the University of Massachusetts at Worcester. "It's able to chop down long RNA strands into small interfering RNAs. That's what Dicer does. In human cells, this enzyme is able to shut down RNAs. That means that the RNA interference phenomenon is working in mammalian cells to cut them down to size. If we didn't have Dicer in human cells, we wouldn't be able to use the newly new antiviral feature of RNA interference. Dicer," Jacque added, "which is actually an RNAase enzyme, came originally from a fruit-fly gene, dcr, and was first described in that insect, Drosophila melanogaster.
"Little is known so far about Dicer," he observed. "It's part of a huge protein complex - probably a million-Dalton aggregate of these molecules - and it recognizes small double-stranded RNAs. A lot of people here at UMass and elsewhere," Jacque noted, "are trying to find out how Dicer works. So far they see a lot of genes and proteins that are part of this enormous conglomerate. Many people are working more specifically on the genetics of this phenomenon. It seems to be a very big complex of proteins that is important for triggering sequence-specific degradation of RNAs. But that's not true just for viruses. It's something that's used for cellular genes. RNA interference might also be used to knock down the gene of a pathogen that is inducing cancer. Which opens a lot more of gene therapy not only for viruses but for many different pathogenic targets."
Jacque, a research-assistant professor at UMass, is first author of a paper in the June 27, 2002, issue of Nature. It's titled: "Modulation of HIV-1 replication by RNA interference."
"We're using Dicer just as a laboratory tool to define targets for the degradation of HIV," Jacque told BioWorld Today.
Basic Science Yes; Gene Therapy For Others
Jacque cited two findings in today's Nature paper: "The first, that we knocked down viral transcription. The second, that we can actually target specifically incoming viral invasion. As soon as the viral genome enters its target T cells, it's marked for degradation by our small RNA duplexes. There's a recent paper in Nature Medicine," he observed, "showing that the HIV virus can be knocked down by RNA interference." (See BioWorld Today, June 10, 2002, p. 1.)
"We are doing basic science," Jacque pointed out. "We are not interested in doing any gene therapy. There are a lot of people around who are specialized on gene therapy. It's my guess that with all these RNA findings coming out, they will jump on it and try to design a new therapeutic approach. RNA interference is so powerful that if they find a way to easily deliver small RNAs into patients, there will certainly be a field for such therapy.
"In our in vitro experiments against HIV," Jacque recounted, "we used isolates from the virus. One approach was to translate two viruses along with the interference RNAs. We got into the T cells with DNA virus. That's the first part of the paper. But the second part is more significant, because we used the whole HIV to infect those cells. So what we had were two copies of RNA in this virus. And early after entry of the virus into the target T cell, it specifically targeted the two copies of the 10,000-base HIV genome. So it was a very early step of infection that enabled us to trigger specific degradation of the genome.
"Because HIV is an RNA virus," Jacque went on, "we thought we could use this feature of RNA interference and see if we could actually knock down virus replication itself. When HIV infects cells, it has to replicate and infect the next cell. It integrates and gets into the latency stage. Then it's just silenced. We thought that using this technique - this unique feature of cells - we could actually knock down replicating cells. If you are able to do that in patients for a long period of time," he pointed out, "you could eventually eradicate that viral cell, because you would just knock it out completely from infected patients. Over a long period of time, treatment with this kind of approach could possibly get the virus out of the patient.
"We used the whole RNA virus. With our labs working on HIV, we had T cells and viral supernatants, so the whole virus is infectious. And we just put that whole virus on top of the target T cells - leading to a productive infection. Before we did that, we introduced these small interference RNAs into those cells, and then infected them. What we looked at was the ability of the virus to infect the cells. When we treated the cells with small interfering RNAs, we knocked down the infection very substantially. We did not let any virus out of the cells, which were actually protected against the infection."
Companion RNA Reports Feature HIV, Polio
In ongoing research, Jacque and his co-authors are "trying to understand a little bit better what's going on in RNA interference and how it works in T cells. Maybe we could find a way to enhance the effects of Dicer in cells. Or activate cells to react against each other by themselves. Very little is known as to how they work in mammalian cells. That's what we're trying to discover right now. Then we can push the cells into this kind of antiviral reaction," Jacque concluded, "using our RNA interference instead of putting the HIV onto the cells."
Today's Nature carries a second - nearly look-alike - article titled: "Short interfering RNA confers intracellular antiviral immunity in human cells." Its senior author is microbiologist Raul Andino from the University of California at San Francisco. Instead of HIV, his paper reports on the polio virus as a model "that may provide a therapeutic strategy against human viruses."