Localization of the secretory granule marker protein chromogranin B in the nucleus. Potential role in transcription control.
Chromogranins A (CGA) and B (CGB) are two major Ca(2+) storage proteins of the secretory granules of neuroendocrine cells. Nevertheless, we found in the present study that CGB was also localized in the nucleus. In immunogold electron microscopy using bovine adrenal medullary chromaffin cells, it was found that the number of CGB-labeled gold particles localized per microm(2) of the nucleus was equivalent to 20% that of CGB-labeled gold particles localized per microm(2) of the secretory granules. Considering that CGB is estimated to exist in the 0.1-0.2-mm range in the secretory granules of bovine chromaffin cells, 20% of these amounts to 20-40 microm. In addition, transfection of CGA and CGB into nonneuroendocrine COS-7 and NIH3T3 cells repeatedly indicated the nuclear localization of CGB in addition to its usual localization in the cytoplasm. Moreover, immunoblot and immunogold electron microscopy analyses of neuroendocrine PC12 cells also showed the existence of endogenous CGB in both the cytosol and the nucleus. Nuclear routing of CGB did not appear to depend entirely upon the nuclear localization signal as some of the nuclear localization signal mutant CGB were still targeted to the nucleus. In gene array assay, CGB was shown to either induce or suppress transcription of many genes including those of transcription factors. Of these we have analyzed eight genes, four induced (zinc finger protein, MEF2C, hCRP2, abLIM) and four suppressed (hcKrox, T3-receptor, troponin C, integrin) using the quantitative reverse transcription-PCR method and spectrophotometry to determine the transcription levels of each mRNA. CGB was shown to increase the transcription of zinc finger protein, MEF2C, hCRP2, and abLIM by 2.5-5-fold while suppressing that of hcKrox, T3-receptor, troponin C, and integrin by 60-75%. Given that MEF2C and hcKrox genes are transcription factors, these results pointed to the transcription control role of CGB in the nucleus.
Yoo
Seung Hyun
SH
National Creative Research Initiative Center for Secretory Granule Research, Korea Advanced Institute of Science and Technology, Yu Sung Gu, Dae Jeon, Korea 305-701. shyoo@kaist.ac.kr
You
Soon Hee
SH
Kang
Moon Kyung
MK
Huh
Yang Hoon
YH
Lee
Choong Sik
CS
Shim
Chan Seob
CS
D016475:3T3 Cells; D000818:Animals; D019556:COS Cells; D002467:Cell Nucleus / Q000648:ultrastructure*; D002522:Chlorocebus aethiops; D053379:Chromogranin A; D053378:Chromogranin B; D002864:Chromogranins / Q000032:analysis* / Q000235:genetics; D003594:Cytoplasmic Granules / Q000648:ultrastructure*; D049452:Green Fluorescent Proteins; D015151:Immunoblotting; D007150:Immunohistochemistry; D008164:Luminescent Proteins / Q000032:analysis / Q000235:genetics; D051379:Mice; D016253:Microscopy, Immunoelectron; D008928:Mitochondria / Q000648:ultrastructure; D016716:PC12 Cells; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection
D053379:Chromogranin A; D053378:Chromogranin B; D002864:Chromogranins; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; C506137:chromogranin B, mouse; C506140:chromogranin B, rat; D049452:Green Fluorescent Proteins