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LIM Kinase Appears Critical in Metastasis, But How?

Robert Finn

Two papers presented at the same poster session of the 40th annual meeting of the American Society for Cell Biology separately concluded that an enzyme named LIM kinase is apparently critical in cell motility and perhaps the process of metastasis. The only problem is that the papers come to opposite conclusions about the enzyme’s effects and suggest opposite strategies for cancer therapy.

The studies conclude that LIM kinase either promotes motility and should be inhibited to arrest metastasis, or the enzyme itself inhibits motility and its effect should be enhanced to arrest metastasis.

In one paper, Kazuyuki Itoh, chief of the Department of Biology at Japan’s Osaka Medical Center for Cancer and Cardiovascular Diseases, demonstrated that human breast cancer cells made to overexpress LIM kinase exhibit increases in adhesion and invasiveness. Itoh and his colleagues found that this was true in both the Matrigel chamber—an in vitro model of metastasis—and when the cells were injected into nude mice.



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Kazuyuki Itoh

 
LIM kinase is part of a signaling cascade that regulates aspects of cell adhesion and movement. It has been known for some time that members of the Rho family of G proteins control certain aspects of adhesion and motility. Among the Rho-protein targets are the ROCK kinases. ROCK kinases phosphorylate and activate LIM kinase, and LIM kinase in turn phosphorylates and activates the protein cofilin. Cofilin is a small protein that depolymerizes actin filaments.



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Kazuyuki Itoh and his colleagues showed that cells that overexpress LIM kinase exhibit increases in adhesion and invasiveness. Human breast cancer cells (MDA-MB-231) overexpressing LIM kinase (bottom row) presented marked polarized focal contacts stained with anti-vinculin antibody (middle column) and enhanced thick stress fiber stained with F-actin (right column) in the cell body, compared with the control vector transfected cells (upper row).

 
Laura M. Machesky, Ph.D., who studies cell motility at the University of Birmingham in the United Kingdom, explained the significance of work on LIM kinase: "We now know that there are only about four or five key proteins that you need to reconstitute motility. One of them is cofilin, which is one of the major players regulating actin disassembly. So if you find an enzyme that regulates cofilin, then you’re regulating major aspects of motility."

In previous work Itoh determined that Y-27632, a specific inhibitor of ROCK kinase, blocked the Rho-mediated effects on the cytoskeleton and reduced the spread of hepatoma cells in rats. The Welfide Corp., Osaka, Japan, from which Itoh receives some funding, is conducting additional research on the anticancer properties of this compound.

But in an interview Itoh pointed out that ROCK has several substrates, and affects not only actin dynamics (through LIM kinase) but also the myosin motor. He is searching for an inhibitor of LIM kinase, which he hopes may more specifically inhibit motility and the process of metastasis.

But another study seems to indicate that LIM kinase itself inhibits motility in neuroblastoma cells. The paper was presented by Gary E. Meyer, a graduate student in the laboratory of Eva L. Feldman, M.D., Ph.D., of the University of Michigan, Ann Arbor. Insulin-like growth factor-I (IGF-I) promotes cell motility, which Meyer measured by observing the tracks etched by neuroblastoma cells in gold-particle-coated coverslips over a 6-hour period. In cells made to overexpress wild-type LIM kinase, the IGF-I–induced increase in motility was blocked.

In an interview Meyer hypothesized, "What we think is going on is that if you’re flooding the cell with too much LIM kinase it’s possible that you’re inactivating so much cofilin that you interfere with the actin recycling process, disrupt the dynamic nature of the lamellipodium [sheetlike protrusions from the leading edge of a moving cell], and thereby inhibit the motility. That’s a model that we’re going to test further."

Itoh, Meyer, and Machesky all recognized the conflicting nature of the two studies, and all three offered explanations that may resolve the apparent discrepancy.

Itoh pointed out that the two laboratories studied LIM kinase in different cell types. "There are a couple of reports which show that this cascade is enhanced in metastatic human cancer, generally speaking. However, we don’t know the exact function of this kinase in normal cells. We are focusing on some very aggressive, metastatic, human cancer cells. In these situations LIM kinase is positively correlated with invasiveness."

Offered Meyer, "One model that you might come up with is that LIM kinase is activated in a particular area of the lamellipodium—like the leading edge— and shutting off cofilin there helps stabilize the branching, whereas cofilin needs to remain active further back in the lamellipodium so it can trim that back end of the meshwork and allow further recycling of the monomers."

And Machesky said, "I don’t have anything that I can say that will resolve the difference between the two papers. I also find it confusing. It may be that the key thing is the balance of cofilin activity so that the cell is functional and can recycle its actin. If you have no cofilin activity you can’t move, and if you have way too much you can’t move either."



             
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