Immunomorphological characterization and effects of 12-(S)-HETE on a dynamic intracellular pool of the alpha IIb beta 3-integrin in melanoma cells.
In metastatic B16a murine melanoma cells, alpha IIb beta 3 integrin was shown to be one of the key adhesion molecules responsible for matrix adhesion and spreading. Upon stimulation, alpha IIb beta 3 can be upregulated at the cell surface due to translocation of the receptor to the plasma membrane from an intracellular pool. Here we have characterized this integrin pool as a tubulovesicular structure (TVS) corresponding to endosomes. TVS was found to be associated temporarily with microtubules and intermediate filaments especially after protein kinase C (PKC) stimulation with a lipoxygenase metabolite of arachidonic acid, 12-(S)-hydroxyeicosatetraenoic acid [12-(S)-HETE]. After PKC stimulation, the predominantly vesicular TVS became elongated and alpha IIb beta 3 appeared at the apical plasma membrane and microvilli. Disruption of either the microtubules or intermediate filaments prevented the 12-(S)-HETE effect both on vesicular to tubular transition of TVS as well as on surface expression of this integrin. The connection with the Golgi system of the integrin-containing TVS was proved by a Golgi-inhibitor (brefeldin A) pretreatment, which prevented the PKC-stimulation-induced TVS elongation and subsequent receptor-upregulation at the cell surface. After a soluble ligand binding (mAb to the alpha IIb beta 3 complex) the surface receptor endocytosed back to the TVS indicating the presence of a dynamic, cytoskeleton associated integrin pool in melanoma cells.
Tímár
J
J
1st Institute of Pathology and Experimental Cancer Research, Semmelweis Medical University, Budapest, Hungary.
Bazaz
R
R
Kimler
V
V
Haddad
M
M
Tang
D G
DG
Robertson
D
D
Tovari
J
J
Taylor
J D
JD
Honn
K V
KV
D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; D000818:Animals; D003599:Cytoskeleton / Q000502:physiology; D011992:Endosomes / Q000648:ultrastructure; D005455:Fluorescent Antibody Technique; D006893:Hydroxyeicosatetraenoic Acids / Q000494:pharmacology*; D016023:Integrins / Q000378:metabolism* / Q000648:ultrastructure; D008546:Melanoma, Experimental; D051379:Mice; D008854:Microscopy, Electron; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D014407:Tumor Cells, Cultured
D006893:Hydroxyeicosatetraenoic Acids; D016023:Integrins; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid