Bradykinin stimulates the ERK-->Elk-1-->Fos/AP-1 pathway in mesangial cells.
Among its diverse biological actions, the vasoactive peptide bradykinin (BK) induces the transcription factor AP-1 and proliferation of mesangial cells (S. S. El-Dahr, S. Dipp, I. V. Yosipiv, and W. H. Baricos. Kidney Int. 50: 1850-1855, 1996). In the present study, we examined the role of protein tyrosine phosphorylation and the mitogen-activated protein kinases, ERK1/2,in mediating BK-induced AP-1 and DNA replication in cultured rat mesangial cells. BK (10(-9) to 10(-7) M) stimulated a rapid increase in tyrosine phosphorylation of multiple proteins with an estimated molecular mass of 120-130, 90-95, and 44-42 kDa. Immunoblots using antibodies specific for ERK or tyrosine-phosphorylated ERK revealed a shifting of p42 ERK2 to a higher molecular weight that correlated temporally with an increase in tyrosine-phosphorylated ERK2. Genistein, a specific tyrosine kinase inhibitor, prevented the phosphorylation of ERK2 by BK. In-gel kinase assays indicated that BK-induced tyrosine phosphorylation of ERK2 is accompanied by fourfold activation of its phosphotransferase activity toward the substrate PHAS-I (P < 0.05). Furthermore, BK stimulated a 2.5-fold increase (P < 0.05) in phosphorylation of Elk-1, a transcription factor required for growth factor-induced c-fos transcription. In accord with the stimulation of Elk-1 phosphorylation, BK induced c-fos gene expression and the production of Fos/AP-1 complexes. In addition, thymidine incorporation into DNA increased twofold (P < 0. 05) following BK stimulation. Each of these effects was blocked by tyrosine kinase inhibition with genistein or herbimycin A. Similarly, antisense oligodeoxynucleotide targeting of ERK1/2 mRNA inhibited BK-stimulated DNA synthesis. In contrast, protein kinase C inhibition or depletion had no effect on BK-induced c-fos mRNA, AP-1-DNA binding activity, or DNA synthesis. Collectively, these data demonstrate that BK activates the ERK-->Elk-1-->AP-1 pathway and that BK mitogenic signaling is critically dependent on protein tyrosine phosphorylation.
El-Dahr
S S
SS
Section of Pediatric Nephrology, Department of Pediatrics, Tulane University School of Medicine, New Orleans, Louisiana 70112, USA.
Dipp
S
S
Baricos
W H
WH
D000818:Animals; D001920:Bradykinin / Q000494:pharmacology*; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D002478:Cells, Cultured; D004247:DNA / Q000096:biosynthesis; D004268:DNA-Binding Proteins; D004789:Enzyme Activation; D005920:Glomerular Mesangium / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D016760:Proto-Oncogene Proteins c-fos / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D018808:Transcription Factor AP-1 / Q000378:metabolism*; D014157:Transcription Factors; D050807:ets-Domain Protein Elk-1
D004268:DNA-Binding Proteins; C494114:Elk1 protein, rat; D011518:Proto-Oncogene Proteins; D016760:Proto-Oncogene Proteins c-fos; D018808:Transcription Factor AP-1; D014157:Transcription Factors; D050807:ets-Domain Protein Elk-1; D019000:Phosphotyrosine; D004247:DNA; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D013755:Tetradecanoylphorbol Acetate; D001920:Bradykinin