For geneticists, interference becomes an asset

A powerful way to learn a gene’s role is to watch how a cell or animal changes when the gene fails. To this end, biologists have used chemicals, X rays, and viruses to introduce mutations.

Their latest trick for disrupting genes is a technique called RNA interference, or RNAi, and a new study offers the first evidence that it works in mammalian cells.

In the February Nature Cell Biology, two investigators from the University of Cambridge in England describe using the new technique to turn off genes in mouse eggs and early embryos.

It “offers many possibilities, both for studying the developmental roles of mouse genes and for using the mouse embryo as a test tube to investigate the functions of ‘housekeeping’ genes, such as those essential for cell division,” says study coauthor Magdalena Zernicka-Goetz.

The RNAi method emerged from puzzlement over so-called antisense RNA (SN: 2/16/91, p. 108). When a gene makes a protein, its DNA spawns an RNA strand, known as messenger RNA (mRNA), that carries the gene’s instructions to a cell’s protein-building factories.

Scientists noticed nearly a decade ago that by adding an RNA strand complementary to the mRNA, they could block protein synthesis. They theorized that such antisense RNA acts by fusing with the mRNA, like a zipper coming together. Investigators also observed, however, that adding sense RNA strands, which are shorter versions of the original mRNA, slowed protein production almost equally well.

It has turned out that most, if not all, of this gene-blocking effect is actually triggered by the double-stranded RNA that researchers inadvertently create when they produce either the sense or antisense strands.

Andrew Fire of the Carnegie Institution of Washington in Baltimore and his colleagues, for example, recently showed that injecting or feeding the worm Caenorhabditis elegans with a gene-specific double-stranded RNA molecule can shut down the gene, even in progeny.

Since it’s easy to create gene-specific double-stranded RNA from a gene’s DNA sequence, interference quickly became a popular way to create mutant worms. Some scientists now plan to use it to block, one by one, each gene in C. elegans. “It’s a very widely used and powerful technology for rapidly testing for the function of a gene,” says Phillip A. Sharp of the Massachusetts Institute of Technology.

RNA interference also works in other worms, plants, fruit flies, and, to some extent, zebrafish, a vertebrate. No one had extended the technique to mammals, however, and some scientists thought that mammalian cells weren’t amenable to it. Since many viruses create double-stranded RNA as they replicate, mammalian cells react strongly to its presence, often by committing suicide.

That didn’t dissuade Zernicka-Goetz and her colleague Florence Wianny. “We had a strong conviction that RNAi had never been thoroughly tested in mammals, especially in embryos,” says Zernicka-Goetz.

Into either immature or fertilized mouse eggs, the pair injected double-stranded RNA segments specific to one of three different genes. In each case, the RNA shut down the protein production of its corresponding gene but not of others.

“I’m really excited. It seems convincing that they’re seeing gene-specific suppression by injection of double-stranded RNA,” says Sharp.

The interference effect wore off after the fertilized eggs went through about six cell divisions.

Consequently, the technique can’t yet create so-called knockout mice, animals in which researchers have permanently disabled a specific gene. Still, John C. Schimenti of the Jackson Laboratory in Bar Harbor, Maine, praises the new research as clearly showing the success of RNA interference in mammals.

“It’s a very exciting result. There’s no doubt people will find ways to put this into action,” the mouse geneticist says.

The work has also opened the eyes of biologists to an intriguing new phenomenon in cells, notes Sharp. The technique seems to exploit a cell’s natural reaction to double-stranded RNA, and scientists are eager to learn more about how—and why—this mysterious gene interference occurs in so many species.

“I think the mechanism is likely to be quite elaborate,” says Sharp.