Regulation of TGF-beta1/MAPK-mediated PAI-1 gene expression by the actin cytoskeleton in human mesangial cells.
The importance of transforming growth factor-beta1 (TGF-beta1) in plasminogen activator inhibitor-1 (PAI-1) gene expression has been established, but the precise intracellular mechanisms are not fully understood. Our hypothesis is that the actin cytoskeleton is involved in TGF-beta1/MAPK-mediated PAI-1 expression in human mesangial cells. Examination of the distributions of actin filaments (F-actin), alpha-actinin, extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) by immunofluorescence and immunoprecipitation revealed that ERK and JNK associate with alpha-actinin along F-actin and that TGF-beta1 stimulation promote the dissociation of ERK and JNK with F-actin. Disassembly of the actin cytoskeleton inhibited phosphorylation of ERK and JNK and modulated PAI-1 expression and promoter activity under both basal and TGF-beta1-stimulated conditions. Stabilizing actin prevented dephosphorylation of ERK and JNK. ERK and JNK inhibitors and overexpressed dominant negative mutants antagonized the ability of TGF-beta1 to increase PAI-1 expression and promoter activity. Disassembly of F-actin also inhibited AP-1 DNA binding activity as determined by electrophoretic mobility shift assay using AP-1 consensus oligonucleotides derived from human PAI-1 promoter. F-actin stabilization prevented loss of AP-1 DNA binding activity. Therefore, changes in actin cytoskeleton modulate the ability of TGF-beta1 to stimulate PAI-1 expression through a mechanism dependent on the activation of MAPK/AP-1 pathways.
Yang
Chen
C
Department of Pathology and Anatomy, Eastern Virginia Medical School, Norfolk, VA 23501, USA. chenyang@mcw.edu
Patel
Keyur
K
Harding
Pamela
P
Sorokin
Andrey
A
Glass
William F
WF
D000199:Actins / Q000502:physiology*; D002460:Cell Line; D003599:Cytoskeleton / Q000502:physiology; D004789:Enzyme Activation; D005786:Gene Expression Regulation; D006801:Humans; D050527:Mesangial Cells / Q000378:metabolism*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D008954:Models, Biological; D017395:Plasminogen Activator Inhibitor 1 / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D051785:Smad Proteins / Q000378:metabolism; D053773:Transforming Growth Factor beta1 / Q000494:pharmacology*
D000199:Actins; D017395:Plasminogen Activator Inhibitor 1; D051785:Smad Proteins; D053773:Transforming Growth Factor beta1; D020928:Mitogen-Activated Protein Kinases