PKC mediates 12(S)-HETE-induced cytoskeletal rearrangement in B16a melanoma cells.
The fatty acid 12(S)-HETE may be a new second messenger capable of activating PKC. In tumor cells 12(S)-HETE stimulates cytoskeleton-dependent cellular responses such as adhesion and spreading. Analysis of 12(S)-HETE effects on B16a melanoma cell cytoskeleton revealed reversible rearrangement of microtubules, microfilaments, the actin-binding proteins, vinculin, myosin heavy (MHC) and light chains (MLC), as well as bundling of vimentin intermediate filaments. The alterations in microfilaments and intermediate filaments occurred very rapidly, i.e., 5 min after exposure of tumor cells to 12(S)-HETE. The 12(S)-HETE-induced cytoskeletal alterations were accompanied by centrifugal organelle-translocation. Interestingly, MLC exhibited clear association with the cytoplasmic organelles. Biochemical analysis of the 12(S)-HETE effect indicated a PKC-mediated reversible hyperphosphorylation of MLC, vimentin, and a 130 kD cytoskeletal-associated protein. Optimal effects were obtained after 5 min treatment with 12(S)-HETE at 0.1 microM concentration. 12(S)-HETE pretreatment induced tumor cell spreading on a fibronectin matrix which required the intactness of all three major cytoskeletal components. The spreading process was dependent upon the activity of PKC. Our data suggest that 12(S)-HETE is a physiological stimulant of PKC. Further, it induces rearrangement of the cytoskeleton of tumor cells in interphase resulting in the stimulation of cytoskeleton-dependent cell activity such as spreading.
Timar
J
J
Department of Radiation Oncology, Wayne State University, Detroit, Michigan 48202.
Tang
D
D
Bazaz
R
R
Haddad
M M
MM
Kimler
V A
VA
Taylor
J D
JD
Honn
K V
KV
D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; D008841:Actin Cytoskeleton / Q000737:chemistry / Q000187:drug effects / Q000648:ultrastructure; D000199:Actins / Q000032:analysis; D000818:Animals; D002118:Calcium / Q000494:pharmacology*; D002465:Cell Movement / Q000502:physiology; D003598:Cytoskeletal Proteins / Q000378:metabolism / Q000502:physiology; D003599:Cytoskeleton / Q000737:chemistry / Q000187:drug effects / Q000648:ultrastructure*; D004305:Dose-Response Relationship, Drug; D005353:Fibronectins / Q000032:analysis; D005455:Fluorescent Antibody Technique; D006893:Hydroxyeicosatetraenoic Acids / Q000494:pharmacology*; D007150:Immunohistochemistry; D007382:Intermediate Filaments / Q000737:chemistry / Q000187:drug effects / Q000648:ultrastructure; D008546:Melanoma, Experimental / Q000737:chemistry / Q000473:pathology* / Q000648:ultrastructure; D051379:Mice; D009218:Myosins / Q000032:analysis; D015388:Organelles / Q000502:physiology / Q000648:ultrastructure; D010743:Phospholipids / Q000494:pharmacology*; D010766:Phosphorylation; D011494:Protein Kinases / Q000494:pharmacology* / Q000502:physiology; D012878:Skin Neoplasms / Q000737:chemistry / Q000473:pathology* / Q000648:ultrastructure; D014407:Tumor Cells, Cultured; D014746:Vimentin / Q000032:analysis; D016596:Vinculin / Q000032:analysis
D000199:Actins; D003598:Cytoskeletal Proteins; D005353:Fibronectins; D006893:Hydroxyeicosatetraenoic Acids; D010743:Phospholipids; D014746:Vimentin; D016596:Vinculin; D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; D011494:Protein Kinases; D009218:Myosins; D002118:Calcium