Active stress kinases in proliferating endothelial cells associated with cytoskeletal structures.
It has become increasingly clear that stress-activated protein kinases have cytoplasmic substrates in addition to well-established transcription factor substrates in cell nuclei. The present study documented specific cytoplasmic locations of these enzymes in proliferating vascular cells. Immunofluorescent staining for active c-jun NH2-terminal kinase (JNK), the precipitation of JNK with microfilaments, and the loss of fiber-associated active JNK after cytochalasin treatment, but not nocodazole treatment, together indicate that active JNK is associated with stress fibers. The lack of complete scaffold colocalization and the total lack of immediate upsteam kinase colocalization along with the inability of JNK inhibitors to alter JNK-microfilament associations suggest that the microfilament association is not simply involved in enzyme activation. In addition, active p38 was found along with vinculin in focal adhesions. Although the p38 in focal adhesions could also be disrupted by cytochalasin treatment, it remained stable after nocodazole treatment. These results support the hypothesis that vascular cell stress kinase enzymes are important for signal transduction in the cytoplasm. The localization of active stress-activated protein kinases to specific cytoskeletal structures in proliferating cells suggests that subsets of these enzymes are involved in signal transduction to and/or from the cytoskeleton under conditions that include vascular cell proliferation.
Hamel
Marianne
M
Department of Biological Sciences, Lehigh University, Bethlehem, Pennsylvania 18015, USA.
Kanyi
Daniela
D
Cipolle
Mark D
MD
Lowe-Krentz
Linda
L
D000313:Adrenal Medulla / Q000098:blood supply; D000818:Animals; D002196:Capillaries / Q000166:cytology; D002417:Cattle; D018929:Cell Culture Techniques; D002460:Cell Line; D049109:Cell Proliferation; D003599:Cytoskeleton / Q000201:enzymology* / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology* / Q000201:enzymology*; D005455:Fluorescent Antibody Technique; D022001:Focal Adhesions / Q000201:enzymology; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism*; D018613:Microscopy, Confocal; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism*; D011494:Protein Kinases / Q000378:metabolism*; D011651:Pulmonary Artery / Q000166:cytology; D016596:Vinculin / Q000378:metabolism; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism*
D016596:Vinculin; D011494:Protein Kinases; D048031:JNK Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D020929:Mitogen-Activated Protein Kinase Kinases