Coupling of the inositol 1,4,5-trisphosphate receptor and chromogranins A and B in secretory granules

The secretory granules of neuroendocrine cells which contain large amounts of Ca2+ and chromogranins have been demonstrated to release Ca2+ in respons e to inositol. 1,4,5 trisphosphate (IP3). Moreover, chromogranin A (CG;A) h as been shown to interact with several secretory granule membrane proteins, including the IP3 receptor (IP3R). To determine whether the IP(3)Rs intera ct directly with chromogranins A and B (CGB), two major proteins of the sec retory granules, we have used purified IP3R from bovine cerebellum in the i nteraction study with CGA and CGB, and have shown that chromogranins A and B directly interact with the IP3R at the intravesicular pH 5.5. Immunogold cytochemical study using the IP3R and CGA antibodies indicated that IP3R-la beled gold particles were localized in the periphery of the secretory granu les, indicating the presence of the IP(3)Rs on the secretory granule membra ne, To determine whether the IP3R and chromogranins A and B are physically linked in the cells, bovine type 1 IP3R (IP3R-1) and CGA or CGB are co-tran sfected into COS-7 cells and co-immunoprecipitation was carried out. Immuno precipitation of the cell extracts demonstrated the presence of CGA-IP3R-1 and CGB-IP3R-1 complexes, respectively, indicating the complex formation be tween the IP3R and chromogranins A and B in native state.