Possible role of protein kinase C in the regulation of intracellular stability of focal adhesion kinase in mouse 3T3 cells.
Effects of various types of protein kinase inhibitor on the adhesion and spreading of BALB/c mouse 3T3 cells and on the phosphorylation and stability of focal adhesion kinase (FAK) in the cells were studied. Inhibitors of protein tyrosine kinases, methyl 2,5-dihydroxycinnamate and herbimycin A, inhibited tyrosine-phosphorylation of FAK and the adhesion of 3T3 cells to fibronectin. Among inhibitors of serine/threonine kinases tested, calphostin C, a specific inhibitor of protein kinase C, inhibited cell spreading rather than cell adhesion, and it induced the decrease of intracellular FAK within 30 min. Inhibitors of tyrosine kinase, A kinase, G kinase, and myosin light chain kinase did not induce such a rapid and specific decrease of FAK. When calphostin C (20 microM) was added to sub-confluent monolayer cultures, serine-phosphorylation of FAK was inhibited by 67% within 2 h, and decrease in the amount of FAK and rounding up of the cells began after 4 h. Label-chase experiments indicated that about 60% of 35S-labeled FAK degraded within 1-2 h after addition of calphostin C to monolayer cultures. These results indicated that serine-phosphorylation of FAK induced by protein kinase C was important in the regulation of metabolic stability of FAK.
Mogi
A
A
Biology Division, National Cancer Center Research Institute, Tokyo, Japan.
Hatai
M
M
Soga
H
H
Takenoshita
S
S
Nagamachi
Y
Y
Fujimoto
J
J
Yamamoto
T
T
Yokota
J
J
Yaoi
Y
Y
D016475:3T3 Cells; D000818:Animals; D016227:Benzoquinones; D002448:Cell Adhesion / Q000187:drug effects / Q000502:physiology; D015815:Cell Adhesion Molecules / Q000378:metabolism*; D002934:Cinnamates / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004795:Enzyme Stability; D005353:Fibronectins; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D007700:Kinetics; D047029:Lactams, Macrocyclic; D051379:Mice; D008807:Mice, Inbred BALB C; D009281:Naphthalenes / Q000494:pharmacology; D010766:Phosphorylation; D010768:Phosphoserine / Q000032:analysis; D019000:Phosphotyrosine / Q000032:analysis / Q000378:metabolism; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D047428:Protein Kinase Inhibitors; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011809:Quinones / Q000494:pharmacology; D017828:Rifabutin / Q000031:analogs & derivatives
D016227:Benzoquinones; D015815:Cell Adhesion Molecules; D002934:Cinnamates; D004791:Enzyme Inhibitors; D005353:Fibronectins; D047029:Lactams, Macrocyclic; D009281:Naphthalenes; D047428:Protein Kinase Inhibitors; D011809:Quinones; D010768:Phosphoserine; D017828:Rifabutin; D019000:Phosphotyrosine; C067472:methyl 2,5-dihydroxycinnamate; C020754:herbimycin; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496051:Ptk2 protein, mouse; D011493:Protein Kinase C; C058819:calphostin C