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Title (en)
Differential Smad2/3 linker phosphorylation is a crosstalk mechanism of Rho/ROCK and canonical TGF-β3 signaling in tenogenic differentiation
Language
English
Description (en)
The transforming growth factor (TGF)-β3 is a well-known inducer for tenogenic differentiation, signaling via the Smad2/3 pathway. Furthermore, other factors like extracellular matrix or mechanical force can induce tenogenic differentiation and possibly alter the response to TGF-β3 by signaling via the Rho/ROCK pathway. The aim of this study was to investigate the interplay of Rho/ROCK and TGF-β3/Smad signaling in tenogenic differentiation, with the Smad2/3 molecule hypothesized as a possible interface. Cultured as monolayers or on collagen I matrices, mesenchymal stromal cells (MSC) were treated with the ROCK inhibitor Y-27632 (10 µM), TGF-β3 (10 ng/ml) or both combined. Control cells were cultured accordingly, without Y-27632 and/or without TGF-β3. At different time points, MSC were analyzed by real-time RT-PCR, immunofluorescence, and Western blot. Cultivation of MSC on collagen matrices and ROCK inhibition supported tenogenic differentiation and fostered the effect of TGF-β3. The phosphorylation of the linker region of Smad2 was reduced by cultivation on collagen matrices, but not by ROCK inhibition. The latter, however, led to increased phosphorylation of the linker region of Smad3. In conclusion, collagen matrices and the Rho/ROCK signaling pathway influence the TGF-β3/Smad2/3 pathway by regulating different phosphorylation sites of the Smad linker region.
Keywords (en)
RHO-Associated Kinases Metabolism; Phosphorylation; Cell Differentiation Drug Effects; Smad2 Protein Metabolism; Smad3 Protein Metabolism; Signal Transduction; Humans; Mesenchymal Stem Cells Metabolism Cytology Drug Effects; Transforming Growth Factor Beta3 Metabolism; Cells, Cultured; Pyridines Pharmacology; Amides Pharmacology; RHO GTP-Binding Proteins Metabolism
DOI
10.1038/s41598-024-60717-z
Author of the digital object
Michaela Melzer  (University of Giessen)
Janina Burk-Luibl  (University of Veterinary Medicine Vienna)
Georgios Scheiner-Bobis  (University of Giessen)
Tilo Pompe  (Leipzig University)
Franziska Ullm  (Leipzig University / Forschungsinstitut für Leder und Kunststoffbahnen)
Sabine Niebert  (University of Veterinary Medicine Vienna)
Manuela Heimann  (University of Giessen)
Format
application/pdf
Size
6.3 MB
Licence Selected
CC BY 4.0 International
Type of publication
Article
Name of Publication (en)
Scientific Reports
Pages or Volume
12
Volume
14
Number
1
Publisher
Nature Portfolio
Publication Date
2024