Harvard Medicine LabWorks

In this video, Tomas Kirchhausen describes the discovery of the traffic-stopping protein dynasore. He and his colleagues are using it in the lab to explore questions such as how membrane traffic sends signals that regulate cell size.

Agent Stops Cell Traffic at Point of Entry

May Serve as Tool for Illuminating Multiple Molecular Pathways

Life at the surface of the cell is anything but placid. Islands of receptor proteins bob on a swirling lipid sea, waiting to attract the attention of a messenger. Once matched, some receptor–messenger pairs disappear as the lipid membrane suddenly invaginates and swallows them.

Endocytosis, this process by which the cell engulfs proteins, microbes, and other molecules, has captured the attention of scientists for hundreds of years.

In 1974, researchers discovered that certain endocytic pathways depend on a remarkable three-legged protein, clathrin.

Over decades, Tomas Kirchhausen, HMS professor of cell biology, and colleagues have learned that clathrin molecules, aided by helper proteins, approach the cell membrane from below and, through an astonishingly swift and graceful sequence of steps, mold it into a bubble-shaped vesicle.

Still the roles of many helpers are poorly defined. One such protein, dynamin, is thought to play an especially important part, coming in at the end and essentially pinching off the completed vesicle. But a clear picture of its comings and goings has been lacking. Eric Macia, Marcello Ehrlich, Ramiro Massol, Kirchhausen, and their colleagues have stopped dynamin in its tracks and now report that the protein plays a dual role: it detaches the completed vesicle from the cell membrane, but it also comes to play earlier in the process, at the point of invagination. Their findings appear in the June 6, 2006 Developmental Cell.

Macia, Ehrlich, and Massol, HMS research fellows in cell biology, working with Kirchhausen and colleagues, screened a library of 16,000 compounds and found one with the ability to block dynamin activity. They added the compound, dynasore, to cultured human cells. Two minutes later, the cells exhibited a complete block of endocytic traffic along the clathrin pathway (see video).

What is more, the endocytic vesicles were frozen in two positions—either fully formed but still attached to the plasma membrane by a small tether or shaped like a U, representing the kinds of half-formed pits one might see just after invagination.

“Dynasore is a cool reagent because you can put it in cells, and within a few minutes, there is a nice block on the entry pathway,” Kirchhausen said.

Cells treated with dynasore rebuffed the advances of a variety of molecules, including transferrin, low-density lipoprotein, and cholera toxin. When the dynamin-blocking agent was washed out, the substances were able to enter.

“This is indeed a terrific tool. Since the compound can rapidly and reversibly block endocytosis, one can do experiments that may not be possible with knockouts or RNAi [RNA interference],” said Venkatesh Murthy, the Morris Kahn associate professor of molecular and cellular biology at Harvard University, who was not an author on the paper.

An even more tantalizing direction would be to use dynasore to keep out certain disease agents, such as cholera toxin. “There is a problem—you would need a way to deliver this to specific cells. You might do that topically,” said Kirchhausen. “In my dreams, I would have a spray with dynasore that I would use to just spritz myself if I had a flu infection. In fact, the influenza virus uses two paths and one of them is dependent on dynamin.”

—Misia Landau