Fluid shear stress-induced TGF-β/ALK5 signaling in renal epithelial cells is modulated by MEK1/2.
Renal tubular epithelial cells are exposed to mechanical forces due to fluid flow shear stress within the lumen of the nephron. These cells respond by activation of mechano-sensors located at the plasma membrane or the primary cilium, having crucial roles in maintenance of cellular homeostasis and signaling. In this paper, we applied fluid shear stress to study TGF-β signaling in renal epithelial cells with and without expression of the Pkd1-gene, encoding a mechano-sensor mutated in polycystic kidney disease. TGF-β signaling modulates cell proliferation, differentiation, apoptosis, and fibrotic deposition, cellular programs that are altered in renal cystic epithelia. SMAD2/3-mediated signaling was activated by fluid flow, both in wild-type and Pkd1-/-cells. This was characterized by phosphorylation and nuclear accumulation of p-SMAD2/3, as well as altered expression of downstream target genes and epithelial-to-mesenchymal transition markers. This response was still present after cilia ablation. An inhibitor of upstream type-I-receptors, ALK4/ALK5/ALK7, as well as TGF-β-neutralizing antibodies effectively blocked SMAD2/3 activity. In contrast, an activin-ligand trap was ineffective, indicating that increased autocrine TGF-β signaling is involved. To study potential involvement of MAPK/ERK signaling, cells were treated with a MEK1/2 inhibitor. Surprisingly, fluid flow-induced expression of most SMAD2/3 targets was further enhanced upon MEK inhibition. We conclude that fluid shear stress induces autocrine TGF-β/ALK5-induced target gene expression in renal epithelial cells, which is partially restrained by MEK1/2-mediated signaling.
Department of Human Genetics, Leiden University Medical Center, 2300 RC, Leiden, The Netherlands.
Steven J
SJ
Kunnen
Department of Human Genetics, Leiden University Medical Center, 2300 RC, Leiden, The Netherlands.
Wouter N
WN
Leonhard
Department of Oral Cell Biology, Academic Centre for Dentistry Amsterdam (ACTA), University of Amsterdam and VU University Amsterdam, 1081 LA, Amsterdam, The Netherlands.
Cor
C
Semeins
Department of Molecular Cell Biology, Cancer Genomics Centre Netherlands, Leiden University Medical Center, 2300 RC, Leiden, The Netherlands.|Department of Gastroenterology-Hepatology, Leiden University Medical Center, 2300 RC, Leiden, The Netherlands.
Lukas J A C
LJAC
Hawinkels
Laboratory for Aero and Hydrodynamics, Delft University of Technology, 2628 CA, Delft, The Netherlands.
Christian
C
Poelma
Department of Molecular Cell Biology, Cancer Genomics Centre Netherlands, Leiden University Medical Center, 2300 RC, Leiden, The Netherlands.
Peter
P
Ten Dijke
Department of Oral Cell Biology, Academic Centre for Dentistry Amsterdam (ACTA), University of Amsterdam and VU University Amsterdam, 1081 LA, Amsterdam, The Netherlands.
Astrid
A
Bakker
Department of Anatomy and Embryology, Leiden University Medical Center, 2300 RC, Leiden, The Netherlands.
Beerend P
BP
Hierck
Department of Human Genetics, Leiden University Medical Center, 2300 RC, Leiden, The Netherlands. d.j.m.peters@lumc.nl.
Dorien J M
DJM
Peters
D028341:Activins / Q000378:metabolism; D000818:Animals; D057134:Antibodies, Neutralizing / Q000494:pharmacology; D015415:Biomarkers / Q000378:metabolism; D002923:Cilia / Q000378:metabolism; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D058750:Epithelial-Mesenchymal Transition / Q000187:drug effects; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D007668:Kidney / Q000166:cytology*; D007687:Kidney Tubules, Proximal / Q000166:cytology; D008024:Ligands; D051379:Mice; D020929:Mitogen-Activated Protein Kinase Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D008954:Models, Biological; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D051381:Rats; D000077293:Receptor, Transforming Growth Factor-beta Type I; D018125:Receptors, Transforming Growth Factor beta / Q000378:metabolism*; D012212:Rheology; D033081:Shear Strength; D015398:Signal Transduction / Q000187:drug effects; D051785:Smad Proteins / Q000378:metabolism; D013314:Stress, Mechanical; D013997:Time Factors; D016212:Transforming Growth Factor beta / Q000378:metabolism
D057134:Antibodies, Neutralizing; D015415:Biomarkers; D008024:Ligands; D018125:Receptors, Transforming Growth Factor beta; D051785:Smad Proteins; D016212:Transforming Growth Factor beta; D028341:Activins; D017346:Protein Serine-Threonine Kinases; D000077293:Receptor, Transforming Growth Factor-beta Type I; C000625841:TGFBR1 protein, human; C000625842:Tgfbr1 protein, mouse; C000625843:Tgfbr1 protein, rat; D020929:Mitogen-Activated Protein Kinase Kinases
Cilia; ERK1/2; Fluid flow; Mechanotransduction; Pkd1 −/−; SMAD2/3 signaling