Kivonat:
Megakaryocytic genes such as alpha IIb beta3 can be expressed by
malignant cells as part of the disturbances in their gene
regulation. However, the function of the gene product greatly
depends on the interaction of the ectopic protein with the new
environment. The outside-in signaling of the ectopically
expressed alpha IIb beta3 integrin was studied in B16a murine
melanoma cells using a monoclonal antibody, specifically
directed to the activated conformation of alpha IIb beta3, PAC-1
and the physiological ligand, fibrinogen. Ligation of alpha IIb
beta3 induced downregulation of FAK but serine phosphorylation
of three protein bands, 20/21, 85 and 140 kDa within 1-15 min.
Flow cytometry indicated that the ligation of the receptor in
B16a cells induces similar to 50%. increase in phosphoserine
positive cells within 5-15 min. 12-lipoxygenase is placed
downstream in the signaling pathway, since ligation of alpha IIb
beta3 induces 12-HETE production within 5 min and pretreatment
of tumor cells with select lipoxygenase inhibitior, Baicalein,
prevents the increase in serine phosphorylation. Confocal
microscopy of adherent tumor cells demonstrated rearrangement of
actin filaments upon alpha IIb beta3 ligation paralleled by
downregulation of p125FAK and phoshotyrosine(+) adhesion plaques
and translocation of PKC alpha to stress fibers and cortical
actin. PKC appears to be the major effector serine kinase of the
alpha IIb beta3-coupled signaling pathway, since pretreatment of
tumor cells with a select PKC inhibitor, Calphostin C, prevents
the ligation-induced serine phosphorylation. Previous studies
have indicated a role for the 12-lipoxygenase-PKC signaling
pathway in platelet aggregation as well as tumor invasion,
therefore the involvement of this cascade in the signaling of
the ectopic alpha IIb beta3 integrin may partially explain its
role in tumor progression.