Abstract
Bone morphogenetic protein (BMP)-1 is expressed by odontoblasts in the dentin-pulp complex. Although the functional effects of BMP-1 on the maturation of various preforms of proteins and enzymes involved in initiating mineralization have been widely observed, how BMP-1 affects cellular molecules remains unknown. We performed a comprehensive analysis of BMP-1-altered glycome profiles and subsequent assays to identify the target glycoproteins in human dental pulp cells (hDPCs) by a glycomic approach. In the presence of BMP-1, a lectin microarray analysis and lectin-probed blotting showed that α2,6-sialylation was significantly attenuated in insoluble fractions from hDPCs. Six proteins were identified by a mass spectrometry analysis of α2,6-sialylated glycoproteins purified using a lectin column. Among them, glucosylceramidase (GBA1) was found to accumulate in the nuclei of hDPCs in the presence of BMP-1. Moreover, BMP-1-induced cellular communication network factor (CCN) 2 expression, which is well known as the osteogenesis/chondrogenesis marker, was significantly suppressed in the cells transfected with GBA1 siRNA. Furthermore, importazole, a potent inhibitor of importin-β-mediated nuclear import significantly suppressed BMP-1-induced GBA1 nuclear accumulation and BMP-1-induced CCN2 mRNA expression, respectively. Thus, BMP-1 facilitates the accumulation of GBA1 in the nucleus through the reduction of α2,6-sialic acid, which potentially contributes to the transcriptional regulation of the CCN2 gene via importin-β-mediated nuclear import pathway in hDPCs. Our results offer new insights into the role of the BMP-1-GBA1-CCN2 axis in the development, tissue remodeling, and pathology of dental/craniofacial diseases.
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Data availability
The mass spectrometry proteomics data have been deposited to the Figshare repository (https://figshare.com/) with the dataset identifier https://doi.org/10.6084/m9.figshare.15128802.
Abbreviations
- BMP-1:
-
Bone morphogenetic protein-1
- hDPCs:
-
Human dental pulp cells
- α2,6-sia:
-
α2,6-Linked sialic acid
- GBA1:
-
Glucosylceramidase
- mTLD:
-
Mammalian tolloid protein
- DMP-1:
-
Dentin matrix protein-1
- DSPP:
-
Dentin sialophosphoprotein
- CCN2:
-
Cellular communication network factor 2
- rhBMP-1:
-
Recombinant human BMP-1
- SNA:
-
Sambucus nigra agglutinin
- SSA:
-
Sambucus sieboldiana agglutinin
- TJA-I:
-
Trichosanthes japonica agglutinin-I
- SDS-PAGE:
-
Sodium dodecyl sulfate-polyacrylamide gel electrophoresis
- CBB:
-
Coomassie brilliant blue
- LAMP1:
-
Lysosome associated membrane protein 1
- DAPI:
-
4′,6-Diamidino-2-phenylindole
- GlcCer:
-
Glucosylceramide
- GBA2:
-
Non-lysosomal β-glucosylceramidase
- GBA3:
-
Cytosolic β-glucosidase
- KLrP:
-
Klotho-related protein
- O-GlcNAc:
-
O-Linked N-acetylglucosamine
- GAPDH:
-
Glyceraldehyde-3-phosphate dehydrogenase
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Acknowledgements
This study was supported by JSPS KAKENHI Grant-in-Aid for Scientific Research (C) #18K09587 and #21K09882. We express our sincere thanks to Dr. Masao YAMADA and GlycoTechnica, Ltd., for conducting the lectin microarray analysis and Shimadzu Techno-Research for conducting the LC-MS/MS analysis. We appreciate Dr. Fumihiko YOSHINO and Dr. Ayaka YOSHIDA for their helpful suggestions. MUROMACHI K expresses a special thanks to Tomomi, Kaho, Reika, and Charo MUROMACHI for their helpful support.
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Muromachi, K., Nakano, R., Fujita-Yoshigaki, J. et al. BMP-1-induced GBA1 nuclear accumulation provokes CCN2 mRNA expression via importin-β-mediated nucleocytoplasmic pathway. J. Cell Commun. Signal. 17, 263–274 (2023). https://doi.org/10.1007/s12079-023-00740-3
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DOI: https://doi.org/10.1007/s12079-023-00740-3