| dc.description.abstracteng | The versatility of intracellular calcium as a second messenger is evident by its ability to mediate such opposing events as neuronal cell growth and apoptosis. One leading hypothesis is that calcium regulates such divergent signaling pathways through the use of functional calcium microdomains. In this thesis, the architectural organization of calcium microdomains is expanded to include accessory proteins which significantly affect the kinetics of calcium signaling. Chromogranin B (CGB) is a calcium binding protein found in secretory granules and the lumen of the endoplasmic reticulum (ER). CGB buffers calcium, and binds to and amplifies the activity of the inositol 1,4,5 trisphosphate receptor (InsP3R). Previous studies have identified two conserved domains in CGB: an N-terminal domain (N-CGB) and a C-terminal domain (C-CGB). N-CGB binds to the third intraluminal loop of the InsP3R and presumably inhibits binding of full-length CGB. This displacement of full-length CGB decreases InsP3R-dependent calcium release, thus altering normal calcium signaling patterns. Here, the role of N-CGB in this process is further characterized, and the heretofore unknown role of C-CGB is identified. Expression of either full-length CGB or C-CGB in cells results in a significant increase in calcium transients. In addition, the calcium signal initiation site in neuronally differentiated PC12 and SHSY5Y cells is affected by C-CGB expression, or altered CGB expression patterns. During the course of these studies, we also found that N-CGB is necessary and sufficient to induce vesicle formation during de novo secretory vesicle biogenesis. Our results strongly suggest that CGB is a part of the InsP3R calcium microdomain, which has numerous regulatory roles, and that CGB plays a critical role in modulating InsP3R-dependent calcium signaling. In fact, dysregulation of CGB has been implicated in the literature in many neurological diseases, consistent with our findings concerning the functional effects of CGB on signal initiation and calcium kinetics in neuronally differentiated cells. Thus, we hypothesize that CGB may be a good candidate for modulating the pathophysiological changes associated with certain neurological diseases. In support of this, we found significant CGB upregulation in the affected brain regions of experimental autoimmune encephalitis (EAE) mice, a mouse model for multiple sclerosis. These results suggest a role for functional CGB-InsP3R calcium microdomains in disease modifying cellular responses. | de |