Pericellular fibronectin is required for RhoA-dependent responses to cyclic strain in fibroblasts.
To test the hypothesis that the pericellular fibronectin matrix is involved in mechanotransduction, we compared the response of normal and fibronectin-deficient mouse fibroblasts to cyclic substrate strain. Normal fibroblasts seeded on vitronectin in fibronectin-depleted medium deposited their own fibronectin matrix. In cultures exposed to cyclic strain, RhoA was activated, actin-stress fibers became more prominent, MAL/MKL1 shuttled to the nucleus, and mRNA encoding tenascin-C was induced. By contrast, these RhoA-dependent responses to cyclic strain were suppressed in fibronectin knockdown or knockout fibroblasts grown under identical conditions. On vitronectin substrate, fibronectin-deficient cells lacked fibrillar adhesions containing alpha5 integrin. However, when fibronectin-deficient fibroblasts were plated on exogenous fibronectin, their defects in adhesions and mechanotransduction were restored. Studies with fragments indicated that both the RGD-synergy site and the adjacent heparin-binding region of fibronectin were required for full activity in mechanotransduction, but not its ability to self-assemble. In contrast to RhoA-mediated responses, activation of Erk1/2 and PKB/Akt by cyclic strain was not affected in fibronectin-deficient cells. Our results indicate that pericellular fibronectin secreted by normal fibroblasts is a necessary component of the strain-sensing machinery. Supporting this hypothesis, induction of cellular tenascin-C by cyclic strain was suppressed by addition of exogenous tenascin-C, which interferes with fibronectin-mediated cell spreading.
Lutz
Roman
R
Friedrich Miescher Institute for Biomedical Research, CH-4056 Basel, Switzerland.
Sakai
Takao
T
Chiquet
Matthias
M
D000199:Actins / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D002448:Cell Adhesion / Q000187:drug effects; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D004622:Embryo, Mammalian / Q000166:cytology; D004789:Enzyme Activation / Q000187:drug effects; D005109:Extracellular Matrix / Q000187:drug effects / Q000378:metabolism; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D005353:Fibronectins / Q000172:deficiency / Q000378:metabolism*; D006493:Heparin / Q000378:metabolism; D039482:Integrin alpha5 / Q000378:metabolism; D051379:Mice; D008840:Microfilament Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D013314:Stress, Mechanical; D019063:Tenascin / Q000235:genetics / Q000378:metabolism; D000070876:Tensins; D015534:Trans-Activators / Q000378:metabolism; D019096:Vitronectin / Q000494:pharmacology; D020742:rhoA GTP-Binding Protein / Q000378:metabolism*
D000199:Actins; D005353:Fibronectins; D039482:Integrin alpha5; D008840:Microfilament Proteins; C511173:Mrtfa protein, mouse; D012333:RNA, Messenger; D019063:Tenascin; D000070876:Tensins; C000603372:Tns1 protein, rat; D015534:Trans-Activators; D019096:Vitronectin; D006493:Heparin; D051057:Proto-Oncogene Proteins c-akt; D048049:Extracellular Signal-Regulated MAP Kinases; D020742:rhoA GTP-Binding Protein