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Scientists shut down genes in poliovirus and HIV in cell cultures


In two new studies, researchers shut down genes in poliovirus and HIV, the virus that causes AIDS. The impaired viruses were often unable to replicate and infect new cells. The findings mark progress in the use of technology to silence genes, a promising but still experimental field of research.

In the poliovirus study, researchers blocked the pathogen early in its life cycle using a technique called RNA interference (RNAi). In this technique, synthetic RNA strands are added to human cells; this triggers a defense mechanism in cells that degrades the corresponding viral RNA and weakens the virus.

Infected cells survive more than a day, but are eventually harmed by emerging siRNA-resistant virus. Bright-field micrographs of cells pre-transfected with buffer, siC and siL, taken 24 hours after infection.

"Our study shows that you can completely protect cultured human cells against infection by the poliovirus," says Raul Andino, of the University of California, San Francisco, who led the study. "The discouraging news is that the virus has found a way to get around RNAi by mutating."

Though they were initially protected against the virus, many cells died a few days later. The surviving viruses, it turned out, had developed mutations that made RNAi ineffective. The technique is extremely specific: Synthetic RNA that differs from its target sequence by a single nucleotide fails to trigger the defense mechanism.

"If a single point mutation can completely inhibit this mechanism, then we have to be very smart in selecting targets," says Andino. Another challenge facing the field is to figure out how best to deliver RNAs to cells. "The method of delivery is the next big question here, and I don't think anybody has a good answer for it," he adds.

In the HIV study, researchers used RNAi to block the virus as soon as it entered human cells. They added short, synthetic strands that matched viral RNA, which caused the degradation of the whole genome. The findings support and expand other recent studies in which RNAi has been used experimentally against HIV.

"We show that you can block the infection at a very early stage and knock down the virus when it gets into the cell," says Jean-Marc Jacque, of the University of Massachusetts Medical School in Worcester.

Inhibition of HIV replication by siRNAs derived from plasmid DNA templates.

Because the virus mutates rapidly, RNAi is unlikely to be useful as a single therapy against HIV, according to Jacque. "In combination with other strategies like protease inhibitors, it may help knock the virus down," he says.

The researchers inhibited the virus in human lymphocytes—cells that are the primary target of HIV. "Knowing that it works against these cells is important because these are the cells that get sick," says Jacque. The findings of both studies were published in Nature online.

See related GNN articles
»Scientists use gene silencing to block hepatitis C virus in mice
»Silencing Genes in HIV

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Gitlin, L. et al. Short interfering RNA confers intracellular antiviral immunity in human cells. Nature. Published online June 26, 2002.
Jacque, J.M. et al. Modulation of HIV-1 replication by RNA interference. Nature. Published online June 26, 2002.

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