Erk1/2 MAPK and caldesmon differentially regulate podosome dynamics in A7r5 vascular smooth muscle cells.
We tested the hypothesis that the MEK/Erk/caldesmon phosphorylation cascade regulates PKC-mediated podosome dynamics in A7r5 cells. We observed the phosphorylation of MEK, Erk and caldesmon, and their translocation to the podosomes upon phorbol dibutyrate (PDBu) stimulation, together with the nuclear translocation of phospho-MEK and phospho-Erk. After MEK inhibition by U0126, Erk translocated to the interconnected actin-rich columns but failed to translocate to the nucleus, suggesting that podosomes served as a site for Erk phosphorylation. The interconnected actin-rich columns in U0126-treated, PDBu-stimulated cells contained alpha-actinin, caldesmon, vinculin, and metalloproteinase-2. Caldesmon and vinculin became integrated with F-actin at the columns, in contrast to their typical location at the ring of podosomes. Live-imaging experiments suggested the growth of these columns from podosomes that were slow to disassemble. The observed modulation of podosome size and life time in A7r5 cells overexpressing wild-type and phosphorylation-deficient caldesmon-GFP mutants in comparison to untransfected cells suggests that caldesmon and caldesmon phosphorylation modulate podosome dynamics in A7r5 cells. These results suggest that Erk1/2 and caldesmon differentially modulate PKC-mediated formation and/or dynamics of podosomes in A7r5 vascular smooth muscle cells.
Gu
Zhizhan
Z
Department of Molecular Pharmacology, Physiology and Biotechnology, Box G-B3, Brown University, Providence, RI 02912, USA.
Kordowska
Jolanta
J
Williams
Geoffrey L
GL
Wang
C-L Albert
CL
Hai
Chi-Ming
CM
D000199:Actins / Q000378:metabolism; D000818:Animals; D002070:Butadienes / Q000494:pharmacology; D002148:Calmodulin-Binding Proteins / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D002273:Carcinogens; D002460:Cell Line; D022081:Cell Surface Extensions / Q000378:metabolism*; D004791:Enzyme Inhibitors; D005786:Gene Expression Regulation; D006160:Guanosine Triphosphate / Q000378:metabolism; D048369:MAP Kinase Kinase 1 / Q000378:metabolism / Q000502:physiology; D020935:MAP Kinase Signaling System; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism* / Q000502:physiology; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000378:metabolism*; D009570:Nitriles / Q000494:pharmacology; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D010766:Phosphorylation; D021381:Protein Transport; D051381:Rats; D014162:Transfection
D000199:Actins; D002070:Butadienes; D002148:Calmodulin-Binding Proteins; D002273:Carcinogens; D004791:Enzyme Inhibitors; D009570:Nitriles; C113580:U 0126; D015240:Phorbol 12,13-Dibutyrate; D006160:Guanosine Triphosphate; D048052:Mitogen-Activated Protein Kinase 3; D048369:MAP Kinase Kinase 1