Scientists at The Scripps Research Institute have described the regulatory mechanism of an important human protein called Rac that controls a number of biological processes and is directly implicated in several human diseases.
Rac is involved in tumor growth and metastasis in cancer; it is important for the proper functioning of immune cells and is necessary for the innate immune response; it is required for neuronal function and has been implicated in neurological diseases and mental retardation.
"Understanding the basic mechanism of how Rac activation is regulated," says Bokoch, "is a key to understanding [these sorts of diseases]."
In an article appearing in the latest issue of the journal Molecular Cell, immunology Professor Gary Bokoch, Ph.D., and his colleagues Céline DerMardirossian and Andreas Schnelzer at Scripps Research have described the molecular mechanism whereby Rac activation is regulated by a molecule called Pak.
The Rac-Pak Connection and Its Relevance to Disease
Rac is one of the most important members of a family of proteins known as the Rho GTPases. This family of proteins binds to a small metabolic product called GTP, which acts as a critical regulator of Rho GTPase activity. This enables Rac to regulate a wide variety of cellular functions that span the entire gamut of a cell's life, from its initial growth and differentiation, to its movement and division, and finally to its death. They are important for gene expression, and they play crucial roles in the ability of innate immune cells to make lethal responses to bacterial infections, of skin cells to cover wounds during the healing process, of vascular cells to make new blood vessels, of cancer cells to metastasize, and of neurons to develop and make proper connections in the brain.
Two years ago Bokoch and his Scripps Research colleagues discovered that Rac is one of the master regulators of cell motility—the molecules driving the process that places the cell's "hands" on the steering wheels and "feet" on the gas pedals. They discovered that Rac is spatially and temporally regulated during leading-edge extension and tail contraction during the movement of human neutrophils—the phagocytic blood cells that chase down, engulf, and destroy bacterial pathogens as part of the body's innate immune response.
One of the big questions that remained unanswered, however, was how Rac was regulated to become active in the first place. What were the master switches that control the activity of Rac and the fundamental cell processes it controls?
All that was known until recently was that inside cells, Rac is controlled by a protein known as RhoGDI, which is in the cell's cytosol. Rac is inactive in resting cells because it is bound to RhoGDI. This keeps the Rac in the cytosol and away from the cellular membrane, where Rac's molecular targets reside. When the cell receives activation signals, the Rac GTPases will dissociate from the RhoGDI in the cytosol and move to the ruffles at the edges of the cell where they are needed. Thus, Rac must be released from RhoGDI for Rac to become active.
"Nobody had any idea how this happened," says DerMardirossian.
Now Bokoch, DerMardirossian, and Schnelzer have discovered the mechanism whereby Rac is released from RhoGDI. In their current study, they show in vitro and in vivo that Rac is released from RhoGDI by an enzyme called p21-activated kinase (Pak). Pak is a kinase enzyme. Its job in the cell is phosphorylation—to attach phosphate groups to other molecules inside the cells in order to modify their function.
Pak attaches its phosphate to two serine residues within the portion of RhoGDI that Rac normally binds to. These bulky and negatively charged phosphates disrupt the normally cozy bond Rac shares with RhoGDI. Freed from the RhoGDI, Rac can then become activated and move about the cell to act on its target molecules and regulate cell function.
Interestingly, one of the targets of active Rac is the enzyme Pak itself. This suggests that Pak and Rac can participate in a positive feedback loop whereby active Rac stimulates Pak, and the active Pak then induces more Rac activation. Such regulation may be important for maintaining continuous cell movement.
The article, "Phosphorylation of RhoGDI by Pak1 mediates dissociation of Rac GTPase" by CŽline DerMardirossian, Andreas Schnelzer, and Gary M. Bokoch appears in the July 2, 2004 issue of the journal Molecular Cell. See http://www.molecule.org.