The role of Jam-C trafficking in endothelial vascular function

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The body's response to a pathogen or tissue injury is essential for survival. White blood cells present in the circulatory system are targeted to the site of injury by signals present on the endothelial cells that line the blood vessels near to the site of injury or infection. This results in rolling of the leukocytes on the endothelium followed by subsequent firm adhesion and crawling before the leukocyte finds a suitable point to pass through the endothelium into the infected tissue. This multi-step cascade is tightly regulated and a number of different protein receptors on both the leukocyte and the endothelium govern each step. A family of proteins present on the endothelial cells (the junctional adhesion molecules or JAMs) have been shown to have important roles in the final step of this inflammatory cascade, the passage of leukocytes through the endothelial cell layer and into the infected tissue. Jam-C, a member of this family, is necessary for this transmigration and has been demonstrated to maintain the directionality of leukocyte movement as well as helping to control the permeability of the endothelial layer. Inhibition of Jam-C function results in leukocytes passing hesitantly through the endothelial cell monolayer and/or returning to the blood flow. Importantly Jam-C expression and function has been associated with multiple inflammatory disease states including, amongst others, arthritis and heart disease. Jam-C is primarily localised to the junctions between endothelial cells where it binds to Jam-C or Jam-B molecules present on neighbouring cells thus helping to create a tight junction. This junction restricts the passage of some molecules and proteins through the endothelial cell layer. When a leukocyte passes through the endothelium the Jam-C present on the endothelial cell is thought to switch to bind a receptor present on the leukocyte surface. Recent research has demonstrated that Jam-C is also present in intracellular vesicular pools and that the amount of Jam-C present at the surface, the cellular junctions and in intracellular vesicles varies following an inflammatory stimulus and this is also true of endothelial cells grown in culture. This indicates that internal pools of Jam-C might be dynamically regulated depending on the stimulus that the cells are exposed to. Internalisation or redistribution of Jam-C might be a means of regulating known Jam-C functions: either by preventing the receptor binding and signalling or by concentrating it in specific pools where it might be used i.e. around transmigrating leukocytes. We propose to develop approaches to monitor the internal trafficking of Jam-C and the cellular machinery required to do this. Often this internalisation and intracellular routing of a surface receptor depends upon amino acid motifs present in the intracellular portion of the receptor. These motifs bind to cellular machinery and determine if the protein is internalised and also what happens subsequently to the now intracellular protein. The receptor might be degraded in the lysosome or the receptor might be recycled back to the cell surface by a number of different routes. Once we have a better mechanistic idea of how Jam-C gets internalised and traffics around the cell we intend to use this information to find novel ways to inhibit this process by knocking out key pieces of machinery or by changing the motifs themselves. This approach should allow us to define the importance of intracellular Jam-C traffic on the known functions of Jam-C. What role does it play in leukocyte migration or in increasing cell permeability? Any new information could translate into new methods for controlling these processes. Such information could eventually be important in controlling inappropriate leukocyte transmigration in chronic inflammatory situations, or in boosting the rate of leukocyte influx in situations where a patient is immune compromised.