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 Ca(2+) and chromogranins have been demonstrated to release Ca(2+) in response to inositol 1,4,5-trisphosphate (IP(3)). Moreover, chromogranin A (CGA) has been shown to interact with several secretory granule membrane proteins, including the IP(3) receptor (IP(3)R). To determine whether the IP(3)Rs interact directly with chromogranins A and B (CGB), two major proteins of the secretory granules, we have used purified IP(3)R from bovine cerebellum in the interaction study with CGA and CGB, and have shown that chromogranins A and B directly interact with the IP(3)R at the intravesicular pH 5.5. Immunogold cytochemical study using the IP(3)R and CGA antibodies indicated that IP(3)R-labeled gold particles were localized in the periphery of the secretory granules, indicating the presence of the IP(3)Rs on the secretory granule membrane. To determine whether the IP(3)R and chromogranins A and B are physically linked in the cells, bovine type 1 IP(3)R (IP(3)R-1) and CGA or CGB are co-transfected into COS-7 cells and co-immunoprecipitation was carried out. Immunoprecipitation of the cell extracts demonstrated the presence of CGA-IP(3)R-1 and CGB-IP(3)R-1 complexes, respectively, indicating the complex formation between the IP(3)R and chromogranins A and B in native state.
Yoo
S H
SH
National Creative Research Initiative Center for Secretory Granule Research, Biomedical Research Center, Korea Advanced Institute of Science and Technology, Yu Sung Gu, Dae Jeon 305-701, Korea. shyoo@cais.kaist.ac.kr
So
S H
SH
Kweon
H S
HS
Lee
J S
JS
Kang
M K
MK
Jeon
C J
CJ
D000818:Animals; D019556:COS Cells; D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels / Q000378:metabolism*; D002417:Cattle; D002531:Cerebellum / Q000737:chemistry; D019439:Chromaffin Cells / Q000378:metabolism*; D053379:Chromogranin A; D002864:Chromogranins / Q000378:metabolism*; D003594:Cytoplasmic Granules / Q000378:metabolism*; D005982:Glutathione Transferase / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007150:Immunohistochemistry; D053496:Inositol 1,4,5-Trisphosphate Receptors; D008854:Microscopy, Electron; D009474:Neurons / Q000737:chemistry; D011233:Precipitin Tests; D011485:Protein Binding; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism*; D014162:Transfection
D015220:Calcium Channels; D053379:Chromogranin A; D002864:Chromogranins; D053496:Inositol 1,4,5-Trisphosphate Receptors; D018160:Receptors, Cytoplasmic and Nuclear; D005982:Glutathione Transferase; D002118:Calcium