Chemical communication between cells keeps tissues functioning and systems coordinated, but eavesdropping on the conversation is challenging. Now, researchers have developed a technique to identify signaling proteins before they leave the cell. The method could help determine which cells are sending which messages—a useful tool for analyzing the interactions occurring in the mixed populations in tissues. One possible application could reveal the cues that control stem cells—an insight that researchers hope could be applied to healing damaged tissues.

The proteins targeted by the method are secreted from one cell and orchestrate the activities of nearby cells. Some signals instruct cells to grow and multiply; others "say" it is time to die. And some signals encourage stem cells—which can mature into a variety of cell types—to differentiate into specific lineages. Understanding cellular signaling is key for biologists hoping to discover how cells respond to one another and their environment. For stem cells in particular, researchers are still puzzling out exactly how they work—for example, molecules from stem cells can heal surrounding tissue. These chemical signals may prompt regeneration of missing cells, recruit other cells to the site or activate some other mechanism.

To study cells, researchers typically culture them in laboratory flasks and dishes. Those cells need a rich stew of proteins, sugars and hormones to stay alive in vitro. The proteins in the culture fluid, however, mask the signals cells send to one another in their natural environment—the body. Cell signals in vivo are even harder to analyze—in any tissue there are many different classes of cells producing a hubbub of different molecular conversations. "We don't know who is talking and who is listening," says Peter Zandstra, a professor of biomedical engineering at the University of Toronto.

Scientists who study cellular communication often solve this signal-to-noise problem by plunking cells into protein-free media for a few hours before analysis. But they have to work fast, because that procedure starves and kills them. Also, those that survive long enough send altered signals.

"It's the classic observer effect," says Balaji Rao, an assistant professor of chemical and bimolecular engineering at North Carolina State University in Raleigh. "By taking away the proteins [in the culture], you've affected the cell physiology."

Rao and his colleagues hit on a conceptually simple solution based on a basic fact from Biology 101: Cells first build and package proteins in specialized internal structures before secreting them. Working with cultured human embryonic stem cells and mouse embryonic fibroblasts (a type of cell that builds the structural framework in tissues and is critical for wound healing), the team isolated the cellular components, or organelles, responsible for assembling the proteins. Those organelles include structures such as the endoplasmic reticulum, the Golgi apparatus and transport vesicles—the latter are essentially bags that ferry the proteins from structure to structure and then out of the cell. Then, using a sophisticated laboratory tool known as a mass spectrometer, they identified the signaling proteins in the isolated organelles, thus acquiring a snapshot of the signaling molecules that would normally be released into the soup of messages outside the cells.

The team, which described the new technique online September 15 in Molecular & Cellular Proteomics, verified its results by comparing the proteins they found with known secretory proteins made by mouse embryonic stem cells and fibroblasts. The technique will be useful for analyzing cells within mixed populations and for tracking how cells respond to a change in their environment, Rao says.

Decoding molecular cues that stem cells send and receive could be key to developing new therapies involving those cells, says Todd McDevitt, an associate professor of biomedical engineering at the Georgia Institute of Technology, who is not involved in the work. For example, discovering a particular protein signal that encourages stem cells to heal damaged tissue could lead to a drug that mimics that protein and enhances tissue repair. A cocktail of such signals might even do away with the need to transplant stem cells for such tasks. This technique will help researchers do the basic research leading to such discoveries, he says. "It could be really transformative."