HHMI physician postdoctoral fellow Michael Yaffe and his colleagues have demonstrated a new mechanism that governs chaperone activity and affects the vitality of cells.

Michael Yaffe's fascination with injury has led him down some unexpected yet fruitful paths. As an HHMI physician postdoctoral fellow at Harvard's Beth Israel Deaconess Medical Center, the critical care physician turned from patching up patients to contemplating how individual cells respond to insult.

Recently, in the course of exploring the role of one type of "molecular chaperone"a protein that acts to get other proteins together and guide them to useful destinationsYaffe and his colleagues demonstrated a new mechanism that governs chaperone activity.

Trained as a surgeon, Yaffe has always been intrigued by the observation that people with seemingly identical injuries often fare quite differently during and after treatment. "I find it hard to look at a patient in the intensive care unit and not wonder about the molecular events transpiring within that person's body," he says. "These patients' bodies are speaking to us in a language we have yet to understand."

One important feature of that lexicon, researchers suspect, is the ability of molecular chaperones to orchestrate the cellular stress response. Since Yaffe was already well versed in the ways of chaperoneshis doctoral research at Case Western Reserve University revealed how a chaperone called TCP-1 assists in the folding of tubulin and actin, two proteins that play a pivotal role in cell divisionhe relished the chance to study them in injured cells. In the laboratory of Lewis Cantley, Yaffe sought to explain how the possible chaperone HSP90 serves in the cell's emergency response team. Soon, however, he was productively diverted from his original plan.

Yaffe began by examining whether HSP90 interacts with signaling molecules called protein kinases, which mediate the cell's response to injury. Interestingly, every protein kinase-HSP90 complex he identified included a third protein: another possible chaperone known as 14-3-3. Several lines of evidence from other laboratories already implicated 14-3-3 as a key regulator of signal molecules involved in a variety of processes, including cell cycle entry and programmed cell death. "I started thinking that 14-3-3 choreographs the dance of cell signaling," Yaffe recalls. "It seems to act like a maypole, around which all the other [protein] players revolve."

Turning his attention to 14-3-3, Yaffe examined how the protein recognizes and binds its many targets. Several of these proteins, when bound to 14-3-3, had been shown to contain the amino acid serine, which bears an attached phosphate groupa sort of removable tag that can be applied whenever 14-3-3's services are needed.

Confirming and extending previous research conducted in the laboratory of Andrey Shaw at Washington University in St. Louis, Yaffe created a "library" of millions of different phosphoserine-studded peptides and tested their ability to bind to 14-3-3. He identified two peptide sequences, or motifs, that bind tightly to the chaperone. Then, working with scientists at London's National Institute for Medical Research, Yaffe determined the crystal structure of 14-3-3 bound to a phosphoserine peptide. Their results appeared in the December 26, 1997, issue of the journal Cell.

Yaffe has consulted protein databases that contain the sequences of thousands of mammalian proteins and found that many of these sequences contain the 14-3-3 binding motif. This suggests that the protein orchestrates a variety of subcellular processes. Meanwhile, however, Yaffe and colleagues at Harvard used his phosphoserine peptide library to reveal another important interaction between the enzyme Pin1 and a group of kinases known to play a crucial role in cell division. That work was published in the December 12, 1997, issue of Science.

Yaffe's research on 14-3-3 has caused him to leave HSP90 behindat least temporarily. Now funded by the National Institutes of Health, he is pursuing several lines of inquiry on his new obsession, the protein he calls "a clever little structure." "Nature," says Yaffe, "seems to do lots of different things with 14-3-3it all depends on where [its binding partners] are phosphorylated."

He's also intrigued by recent reports from other laboratories that implicate 14-3-3's involvement in the cellular response to injury following carotid artery trauma and irradiation. Yaffe didn't plan it that way, but his detour may bring him full circleand back to studying injured cells in sick patients.

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