Sulphation of secreted phosphoprotein I (SPPI, osteopontin) is associated with mineralized tissue formation
Nagata, T.; Todescan, R.; Goldberg, H.A.; Zhang, Q.; Sodek, J.
Biochemical and Biophysical Research Communications 165(1): 234-240
1989
ISSN/ISBN: 0006-291X
PMID: 2686645
Accession: 007845147
Secreted phosphoprotein I (SPPI) is a prominent structural protein in mineralized connective tissues. Rat bone cells in culture produce several forms of SPPI that differ in post-translational modifications such as phosphorylation and sulphation. To determine the significance of protein sulphation in bone formation, the synthesis of SPPI was studied in vitro using rate bone marrow cells (RBMC) which form bone-like tissue when grown in the presence of dexamethasone (Dex) and .beta.-glycerophosphate (.beta.-GP). In the presence of 10-7 M Dex SPPI expression was stimulated 4-5-fold. Radiolabeling multilayered RBMCs for 48 h with [35S]-methionine, Na2[35SO4], or Na3[32PO4] revealed that two major phosphorylated forms of SPPI were secreted into the culture medium: a highly phosphorylated form migrating at 44 kDa on 15% SDS-PAGE and a less phosphorylated 55 kDa form. In the mineralized tissue formed in the presence of Dex and .beta.-GP, both forms of SPPI, in addition to proteoglycans and a 67 kDa protein, incorporated significant amounts of [35SO4]. Sulphation of SPPI was not observed in the absence of mineral formation, indicating that the sulphation of SPPI is closely associated with mineralization and that it can be used as a sensitive and specific marker for the osteoblastic phenotype.