Cooper's lab and others also had found several proteins that regulate Arp2/3 activity. In yeast, whose complete genetic code has been sequenced and can be searched, there appears to be a total of six proteins that activate or deactivate Arp2/3. These proteins are regularly found in actin patches on the surfaces of yeast cells, which are areas where endocytosis is about to take place.
"We wanted to know why there needed to be six of these proteins, if any one could either turn Arp2/3 on or off," says Galletta.
To learn more, Galletta mutated each of the six proteins alone and in various combinations. Researchers also attached two different fluorescent labels to other proteins involved in the early and late stages of endocytosis. The labels let them use a light microscope to make movies of hundreds of actin patches in the various yeast lines, recording the patches' movement across cell membranes and into cells.
With the help of a computerized tracking program from Anders Carlsson, Ph.D., professor of physics at Washington University, they quantitatively analyzed the results, uncovering both dramatic and subtle variations in actin patch activity caused by the mutations.
"We found that sometimes the Arp2/3 regulatory proteins had a unique function of their own, and sometimes they cooperated," Galletta says. "That was surprising, because in the test tube, all of these proteins have very similar functions."
In addition to actin networks' use by immune cells chasing invading microorganisms, those same microorganisms sometimes hijack the actin networks in host cells, effectively seizing the cells' steering wheels and driving them around. The networks also play important roles in the developing embryo, enabling cells to assume specialized shapes or migrate to where they need to be in the body.
|Contact: Michael C. Purdy|
Washington University School of Medicine