While completing her graduate work on mouse embryonic development, Karen Liu became frustrated with the limitations in the methodology available to her. She decided that she needed new tools that would enable her to study more precisely the timing of events that occur during embryonic development.

After completing her PhD, Liu joined the laboratory of molecular biologist Gerald Crabtree at Stanford University, under the co-mentorship of craniofacial surgeon Michael Longaker. Combining her mentors' expertise with her own, Liu upgraded her developmental biology toolbox and has used the new tools to prevent cleft palate in a mouse genetic model of the condition (see page 79).

The Crabtree lab had developed an 89-amino-acid tag, called FRB*, which renders any protein fused with it unstable. When the drug rapamycin binds to the tag, however, it stabilizes the protein, restoring its function. Liu decided to use this technique to study development in vivo. “I was lucky to have arrived at a time when these tools were available and I could move them in the direction I wanted,” she explains.

To test the technique, the Crabtree lab had already constructed mice containing FRB*-tagged glycogen synthase kinase (GSK-3β). This is a component of many important developmental signalling pathways and is involved in several diseases. The researchers had shown that the mice produced an unstable GSK-3β protein. Liu reasoned that the mice would therefore have a similar phenotype as conventional 'knockout' mice lacking GSK-3β. To test this hypothesis she first looked at GSK-3β knockouts and found that these mice were born with defective fusion of the palate (cleft palate) and of the sternum, or breastbone. Exactly the same defects appeared in her FRB*–GSKβ mice without rapamycin, confirming that the tagged GSK-3β was not functional.

The final test came when Liu and her colleagues gave rapamycin injections to pregnant mice carrying rapamycin-responsive mutant embryos. The mutant fetuses developed with a normal palate. “I thought we did not have any mutants in the litter,” says Liu. But genetic analysis revealed that the newborn mice did indeed carry the mutant GSK-3β. “I did not believe it,” she laughs. “We repeated the genotyping and then repeated the experiment several times.”

Rapamycin given at an early stage of development prevented cleft palate; at a later stage it prevented the defect in the sternum. This indicated two distinct windows of time when GSK-3β is needed in development.

“The concept of using a small molecule such as rapamycin to correct a birth defect is something I could have only dreamed of 20 years ago,” says Longaker, who is a co-author on the paper. This experiment is, however, a long way away from thinking about a similar approach to therapy, he cautions.

“We were not sure it would work,” says Liu. “It was not clear we could deliver the drug to mice, let alone pregnant ones.” Having a team of scientists with diverse backgrounds helped make the study possible, says Longaker. “It would have been hard to do in any one lab,” he says. “An interdisciplinary team needs a champion and Karen was the glue that kept the project together.”

Liu has just set up her own laboratory at King's College in London and plans to continue with the project and refine the FRB* tag system. She plans to develop different combinations of tags and drugs and also to create mice with tissue-specific mutations that can be 'rescued' using this system.