Adhesion of metastatic, ras-transformed NIH 3T3 cells to osteopontin, fibronectin, and laminin.
We previously reported that H-ras-induced metastatic ability in murine NIH 3T3 cells is accompanied by increased expression of osteopontin (OPN). OPN is a secreted phosphoprotein that contains a GRGDS amino acid sequence, suggesting adhesive function, but the function of OPN in tumor cells remains poorly understood. Here we report that PAP2 cells (ras-transformed, metastatic NIH 3T3 cells) adhere and spread on OPN-coated substrates, while NIH 3T3 cells adhere and spread poorly on OPN. A similar pattern was seen for adhesion to laminin, while both cell lines adhered equally well to fibronectin. Adhesive interactions to OPN, laminin, and fibronectin were specific and were blocked by GRGDS (but not control GRGESP) peptides. The kinetics of adhesion to all three substrates was examined. Maximum adhesion was observed at 30-60 min, with reduced adhesion thereafter. We also purified metabolically labeled [32P]OPN secreted by PAP2 cells. Labeled OPN bound better in solution to PAP2 cells than to NIH 3T3 cells, and binding to both cell lines was blocked by GRGDS peptides, results that are consistent with the adhesion and spreading of these cells to OPN-coated substrates. Malignant PAP2 cells thus not only secrete increased levels of OPN, relative to NIH 3T3 cells, but also adhere better to this protein. While the target of OPN secreted by tumor cells is not known, our results raise the possibility that tumor cells that secrete OPN may also bind this protein and that this binding may function in autocrine-type signal transduction important to malignancy.
Chambers
A F
AF
London Regional Cancer Centre, Department of Oncology, University of Western Ontario, Canada.
Hota
C
C
Prince
C W
CW
D016475:3T3 Cells / Q000378:metabolism / Q000473:pathology* / Q000502:physiology; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002448:Cell Adhesion / Q000502:physiology; D015815:Cell Adhesion Molecules / Q000378:metabolism / Q000502:physiology; D002471:Cell Transformation, Neoplastic / Q000235:genetics / Q000473:pathology*; D005353:Fibronectins / Q000378:metabolism / Q000502:physiology*; D011905:Genes, ras / Q000502:physiology*; D007797:Laminin / Q000378:metabolism / Q000502:physiology*; D051379:Mice; D008969:Molecular Sequence Data; D009362:Neoplasm Metastasis; D009842:Oligopeptides / Q000378:metabolism; D053495:Osteopontin; D010761:Phosphorus Radioisotopes; D011485:Protein Binding; D012680:Sensitivity and Specificity; D012795:Sialoglycoproteins / Q000378:metabolism / Q000502:physiology*
D015815:Cell Adhesion Molecules; D005353:Fibronectins; D007797:Laminin; D009842:Oligopeptides; D010761:Phosphorus Radioisotopes; D012795:Sialoglycoproteins; C506896:Spp1 protein, mouse; D053495:Osteopontin; C047981:arginyl-glycyl-aspartic acid