This project will focus on the intracellular traffic of plasma membrane constituents in acinar cells isolated from rabbit lacrimal glands. The lacrimal glands are effector organs in a servomechanism that helps maintain the health of the ocular surface, and insufficient lacrimal function leads to painful dry eye conditions. Lacrimal insufficiency can result from hormonal perturbations or from local autoimmune phenomena. It is particularly urgent that the mechanisms leading to autoimmune lacrimal insufficiency be understood. This phenomenon is often accompanied by autoimmune salivary gland dysfunction, with serious consequences for oral health, and by numerous systemic disorders triggered, at least in part, by immune complex deposition. these symptoms characterize Sjogren's Syndrome, which affects more than one million American women over age 55. Recent subcellular fractionation and morphological studies indicate that Na,K-ATPase and other acinar cell plasma membrane proteins participate in an extensive recycling traffic to and from an intracellular pool, which appears to be organized into early and late endocytic compartments. Segregation mechanism appear to permit fluid phase traffic between the two compartments but allow Na,K-ATPase to travel from the late to the early compartment only in response to cholinergic stimulation. Excessive stimulation appears to alter the microdomain organization of the late compartment and to impair its ability to participate in intracellular fluid phase traffic. The existence of such an extensive, secretagogue- controlled intracellular traffic suggests a working hypothesis in which acinar cells trigger local autoimmune reactions by functioning analogously to specialized antigen-presenting cells. Preliminary data confirm that acinar cells contain a potentially immunogenic protein (LG- Ag2) and that they can express several components of the machinery for antigen presentation, including cathepsin B, which might excise immunogenic peptide fragments, and Class II histocompatibility molecules, which might bind immunogenic fragments and display them to T cells. It is also possible that additional peptides occurring in the lacrimal interstitium can provide accessory signals for T cell activation.
The specific aims of the proposed studies are to: 1) Delineate and survey the microdomain organizations of the early and late endocytic compartments. 2) Identify mechanisms responsible for re-organizing the late endocytic compartment following optimal and excessive cholinergic stimulation. 3) Map the subcellular distributions of cathepsins B and D, LG-Ag2, and Class II histocompatibility molecules to determine whether they can accumulate in compartments that communicate with the plasma membrane. If the hypotheses explored by the project are proven correct, then progress will have been made in the understanding of events underlying insertion and retrieval of membrane transporters, and the stage will be set for attempts to understand T cell activation by acinar cells.
