Osteopontin increases CD44 expression and cell adhesion in RAW 264.7 murine leukemia cells.
BACKGROUND
Osteopontin (OPN) is an inducible cell attachment protein which binds alphavbeta3-integrin and CD44 receptors to promote tumor metastasis. We hypothesized that OPN alters expression of its CD44 receptor to promote neoplastic cell migration.


METHODS
RAW264.7 cells were stimulated with OPN (0-10 nM) for 0-12 hours to determine the time- and concentration-dependence of CD44 protein and mRNA expression. In selected instances, a competitive ligand for the alphavbeta3-integrin, GRGDSP (50 nM), or an inhibitor of protein synthesis, anisomycin (10 microg/ml), was added. Cell adhesion to hyaluronan was assayed with the crystal violet assay.


RESULTS
OPN upregulates plasma membrane total CD44 protein in a concentration-(ANOVA P = 0.001) and time-dependent (ANOVA P = 0.001) fashion. CD44v6 is not altered. Cell adhesion to hyaluronate increases in parallel with CD44 expression. Steady state mRNA levels for CD44 are not altered by OPN. 5 nM OPN increases CD44 protein half-life from 105 +/- 11 minutes to 278 +/- 15 minutes. (P < 0.03) Blockade of either alphavbeta3-integrin ablates the OPN-dependent increase in CD44.


CONCLUSIONS
These data indicate that OPN increases plasma membrane CD44 expression and cell adhesion by binding to its alphavbeta3-integrin receptor. We conclude that OPN may promote tumor metastatic behavior by CD44 expression.
Department of Surgery, Duke University Medical Center, 110 Bell Bldg., Box 3522, Durham, NC 27710, USA.
Carlos E
CE
Marroquin
Laura
L
Downey
Hongtao
H
Guo
Paul C
PC
Kuo
D000818:Animals; D000841:Anisomycin / Q000494:pharmacology; D002448:Cell Adhesion; D045744:Cell Line, Tumor; D018960:Hyaluronan Receptors / Q000235:genetics / Q000378:metabolism*; D007938:Leukemia / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D051379:Mice; D009374:Neoplasms, Experimental / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D009842:Oligopeptides / Q000494:pharmacology; D053495:Osteopontin; D012333:RNA, Messenger / Q000378:metabolism; D012795:Sialoglycoproteins / Q000378:metabolism / Q000494:pharmacology*
D018960:Hyaluronan Receptors; D009842:Oligopeptides; D012333:RNA, Messenger; D012795:Sialoglycoproteins; C506896:Spp1 protein, mouse; D053495:Osteopontin; D000841:Anisomycin; C058161:glycyl-arginyl-glycyl-aspartyl-seryl-proline