Possible role of protein kinase C in the regulation of intracellular stability of focal adhesion kinase in mouse 3T3 cells

• Published in: FEBS letters 1995-10, Vol.373 (2), p.135-140
• Main Authors: Mogi, Akira, Hatai, Mika, Soga, Hisae, Takenoshita, Seiichi, Nagamachi, Yukio, Fujimoto, Jiro, Yamamoto, Tadashi, Yokota, Jun, Yaoi, Yoshihito
• Format: Article
• Language: English
• Subjects: 2,5-MeC, methyl-2,5-dihydroxycinnamate; 3T3 Cells; Animals; Benzoquinones; Calphostin C; Cell Adhesion - drug effects; Cell Adhesion - physiology; Cell Adhesion Molecules - metabolism; Cinnamates - pharmacology; Enzyme Inhibitors - pharmacology; Enzyme Stability; FAK; FAK, pp125125, focal adhesion kinase; Fibronectin; Fibronectins; Focal adhesion kinase; Focal Adhesion Kinase 1; Focal Adhesion Protein-Tyrosine Kinases; Kinetics; Lactams, Macrocyclic; Mice; Mice, Inbred BALB C; Naphthalenes - pharmacology; Phosphorylation; Phosphoserine - analysis; Phosphotyrosine - analysis; Phosphotyrosine - metabolism; PKC, protein kinase C; Protein kinase C; Protein Kinase C - antagonists & inhibitors; Protein Kinase C - metabolism; Protein Kinase Inhibitors; Protein-Tyrosine Kinases - antagonists & inhibitors; Protein-Tyrosine Kinases - metabolism; Quinones - pharmacology; Rifabutin - analogs & derivatives; SDS-PAGE, sodium dodecyl sulfate-polyacrylamide gel electrophoresis
• ISSN: 0014-5793; 1873-3468
• DOI: 10.1016/0014-5793(95)01014-6

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 μM) 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.