Stimulation of pro-alpha(1)(I) collagen by TGF-beta(1) in mesangial cells: role of the p38 MAPK pathway.
Transforming growth factor-beta(1) (TGF-beta(1)) is a potent inducer of extracellular matrix protein synthesis and a key mediator of renal fibrosis. However, the intracellular signaling mechanisms by which TGF-beta(1) stimulates this process remain incompletely understood. In this report, we examined the role of a major stress-activated intracellular signaling cascade, belonging to the mitogen-activated protein kinase (MAPK) superfamily, in mediating TGF-beta(1) responses in rat glomerular mesangial cells, using dominant-negative inhibition of TGF-beta(1) signaling receptors. We first stably transfected rat glomerular mesangial cells with a kinase-deleted mutant TGF-beta type II receptor (TbetaR-II(M)) designed to inhibit TGF-beta(1) signaling in a dominant-negative fashion. Next, expression of TbetaR-II(M) mRNA was confirmed by Northern analysis. Cell surface expression and ligand binding of TbetaR-II(M) protein were demonstrated by affinity cross-linking with (125)I-labeled-TGF-beta(1). TGF-beta(1) rapidly induced p38 MAPK phosphorylation in wild-type and empty vector (pcDNA3)-transfected control mesangial cells. Interestingly, transfection with dominant-negative TbetaR-II(M) failed to block TGF-beta(1)-induced p38 MAPK phosphorylation. Moreover, dominant-negative TbetaR-II(M) failed to block TGF-beta(1)-stimulated pro-alpha(1)(I) collagen mRNA expression and cellular protein synthesis, whereas TGF-beta(1)-induced extracellular signal-regulated kinase (ERK) 1/ERK2 activation and antiproliferative responses were blocked by TbetaR-II(M). In the presence of a specific inhibitor of p38 MAPK, SB-203580, TGF-beta(1) was unable to stimulate pro-alpha(1)(I) collagen mRNA expression in the control and TbetaR-II(M)-transfected mesangial cells. Finally, we confirmed that both p38 MAPK activation and pro-alpha(1)(I) collagen stimulation were TGF-beta(1) effects that were abrogated by dominant-negative inhibition of TGF-beta type I receptor. Thus we show first demonstration of p38 MAPK activation by TGF-beta(1) in mesangial cells, and, given the rapid kinetics, this TGF-beta(1) effect is likely a direct one. Furthermore, our findings suggest that the p38 MAPK pathway functions as a component in the signaling of pro-alpha(1)(I) collagen induction by TGF-beta(1) in mesangial cells.
Chin
B Y
BY
Toxicological Sciences, Environmental Health Sciences, The Johns Hopkins Medical Institutions, Baltimore, Maryland 21205, USA.
Mohsenin
A
A
Li
S X
SX
Choi
A M
AM
Choi
M E
ME
D000818:Animals; D002478:Cells, Cultured; D003094:Collagen / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D024042:Collagen Type I; D000090602:Collagen Type I, alpha 1 Chain; D004789:Enzyme Activation; D005799:Genes, Dominant; D005920:Glomerular Mesangium / Q000166:cytology / Q000378:metabolism*; D007930:Leucine / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism / Q000502:physiology*; D010766:Phosphorylation; D011498:Protein Precursors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D018125:Receptors, Transforming Growth Factor beta / Q000235:genetics; D011994:Recombinant Proteins / Q000494:pharmacology; D013936:Thymidine / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000494:pharmacology*; D053773:Transforming Growth Factor beta1; D048051:p38 Mitogen-Activated Protein Kinases
D024042:Collagen Type I; D000090602:Collagen Type I, alpha 1 Chain; D011498:Protein Precursors; D012333:RNA, Messenger; D018125:Receptors, Transforming Growth Factor beta; D011994:Recombinant Proteins; C508962:Tgfb1 protein, rat; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; D003094:Collagen; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D007930:Leucine; D013936:Thymidine