PKCalpha translocation is microtubule-dependent in passaged smooth muscle cells.
The translocation of protein kinase C (PKC) isozymes from their inactive cell locus to a variety of cytoskeletal, organelle, and plasmalemmal sites is thought to play an important role in their activation and substrate specificity. We have utilized confocal microscopy to compare phorbol 12, 13 dibutyrate (PDB) - stimulated translocation of PKCalpha in cultured cells derived from rat vascular smooth muscle. In enzymatically dispersed, passaged smooth muscle cells, PKCalpha was uniformly distributed throughout the unstimulated cell. PDB stimulation resulted in extensive association of the PKCalpha into filamentous strands with subsequent accumulation of the isoform in the peri-nuclear region of the cell. Dual immunostaining indicated that PKCalpha was extensively colocalized with microtubules in the interval immediately following PDB stimulation but was largely disassociated from microtubules at 10 min, at which time the translocation of PKCalpha to the peri-nucleus/nucleus was nearly complete. It was further found that the use of colchicine to disrupt the microtubules caused the loss of PKCalpha translocation to the peri-nuclear region. By comparison, cytochalasin B disruption of actin microfilaments had no significant effect on this parameter. The data suggest that PDB stimulation results in a transient association of PKCalpha with cell microtubules and that the microtubules play an important role in the translocation of PKCalpha from the cytosol in passaged cells derived from rat aortic smooth muscle.
Battistella-Patterson
A S
AS
Department of Physiology, Marshall University School of Medicine, Huntington, WV 25704, USA.
Fultz
M E
ME
Li
C
C
Geng
W
W
Norton
M
M
Wright
G L
GL
D008841:Actin Cytoskeleton / Q000187:drug effects / Q000378:metabolism; D000818:Animals; D001011:Aorta / Q000166:cytology; D001692:Biological Transport / Q000502:physiology; D002478:Cells, Cultured; D003078:Colchicine / Q000494:pharmacology; D003571:Cytochalasin B / Q000494:pharmacology; D003600:Cytosol / Q000378:metabolism; D007527:Isoenzymes / Q000378:metabolism*; D008297:Male; D008870:Microtubules / Q000187:drug effects / Q000378:metabolism*; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000201:enzymology*; D011493:Protein Kinase C / Q000378:metabolism*; D051571:Protein Kinase C-alpha; D051381:Rats; D017207:Rats, Sprague-Dawley
D007527:Isoenzymes; D003571:Cytochalasin B; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D003078:Colchicine