Phosphorylated Osteopontin Secreted from Cancer Cells Induces Cancer Cell Motility

Abstract

Osteopontin (OPN) plays a pivotal role in cancer cell invasion and metastasis. Although OPN has a large number of phosphorylation sites, the functional significance of OPN phosphorylation in cancer cell motility remains unclear. In this study, we attempted to investigate whether phosphorylated OPN secreted from cancer cells affect cancer cell migration. Quantitative PCR and Western blot analyses revealed that MDA-MB435S, A549, and H460 cells highly expressed OPN, whereas the OPN expression levels in H358, MIAPaca-2, and Panc-1 cells were quite low or were not detected. Compared with the cancer cell lines with a low OPN expression, the high OPN-expressing cancer cell lines displayed a higher cell migration, and the cell migration was suppressed by the anti-OPN antibody. This was confirmed by the OPN overexpression in H358 cancer cells with a low endogenous OPN. Phos-tag ELISA showed that phosphorylated OPN was abundant in the cell culture media of A549 and H460 cells, but not in those of MDA-MB435S cells. Moreover, the A549 and H460 cell culture media, as well as the MDA-MB435S cell culture media with a kinase treatment increased cancer cell motility, both of which were abrogated by phosphatase treatment or anti-OPN antibodies. These results suggest that phosphorylated OPN secreted from cancer cells regulates cancer cell motility.

Osteopontin/secreted phosphoprotein‐1 harnesses glial‐, immune‐, and neuronal cell ligand‐receptor interactions to sense and regulate acute and chronic neuroinflammation

“… 47 , 48 The highly acidic charge of OPN/SPP1 confers on the bone's ability to regulate mineralization in extracellular matrix (ECM). 47 , 49 N‐linked and O‐linked glycosylation 50 , 51 , 52 as well as transglutamination, which results in the increased polymerization of OPN/SPP1 in the ECM, can also play a role in either masking interaction domains or enhancing select functions of OPN/SPP1 in the particular cell and tissue relevant context. 53 , 54 , 55 , 56 Given the sheer number of acidic residues and, therefore, the limits of studying the direct contributions of phosphorylation to OPN/SPP1 activity in vivo, most studies in this regard are biochemical, use modified peptides, mass spectrometry analyses, and cell‐based assays to then infer the likely impact of phosphorylation on functions like integrin‐binding ability.…”

Proteomics, Phosphoproteomics and Mirna Analysis of Circulating Extracellular Vesicles through Automated and High-Throughput Isolation

“…Previous studies indicated that secreted phosphoprotein 1 (SPP1) was involved in the recurrence and metastasis of prostate cancer [46] and regarded as a potential biomarker and therapeutic target for metastatic castration-resistant prostate cancer (mCRPC) [47]. Meanwhile, it had been shown that phosphorylated SPP1 secreted from cancer cells regulated cancer cell motility in human lung cancer [48]. In our data, we also found significant upregulation of SPP1 in the prostate cancer group (Figure 5E), and phosphorylation levels of SPP1 at 219 sites in the prostate cancer group were significantly higher than that in controls (Figure 5H).…”

Osteopontin: An important protein in the formation of kidney stones

“…OPN is phosphorylated by Golgi-casein kinases, and the phosphorylation level and site are affected by the o-glycosylation state (OPN with five o-glycosylation sites deleted from threonine/proline rich regions increases cell adhesion activity and phosphorylation, inhibits integrin association) ( Kariya et al, 2014 ) and 1,25-(OH)2D3 (induced upregulation of OPN mRNA expression, while promoting phosphorylated OPN converts to the non-phosphorylated state) ( Safran et al, 1998 ). The degree of phosphorylation of OPN affects cell adhesion and interaction with other proteins ( Kariya et al, 2021 ).…”