Increased cAMP levels modulate transforming growth factor-beta/Smad-induced expression of extracellular matrix components and other key fibroblast effector functions.
cAMP is a key messenger of many hormones and neuropeptides, some of which modulate the composition of extracellular matrix. Treatment of human dermal fibroblasts with dibutyryl cyclic AMP and forskolin antagonized the inductive effects of transforming growth factor-beta (TGF-beta) on the expression of collagen, connective tissue growth factor, tissue inhibitor of matrix metalloproteinase-1, and plasminogen activator inhibitor type I, four prototypical TGF-beta-responsive genes. Increased intracellular cAMP prevented TGF-beta-induced Smad-specific gene transactivation, although TGF-beta-mediated Smad phosphorylation and nuclear translocation remained unaffected. However, increased cAMP levels abolished TGF-beta-induced interaction of Smad3 with its transcriptional co-activator cAMP-response element-binding protein (CREB)-binding protein (CBP)/p300. Overexpression of the transcriptional co-activator CBP/p300 rescued Smad-specific gene transcription in the presence of cAMP suggesting that sequestration of limited amounts of CBP/p300 by the activated cAMP/CREB pathway is the molecular basis of this inhibitory effect. These findings were extended by two functional assays. Increased intracellular cAMP levels suppressed the inductive activity of TGF-beta to contract mechanically unloaded collagen lattices and resulted in an attenuation of fibroblast migration of mechanically induced cell layer wounds. Of note, cAMP and TGF-beta synergistically induced hyaluronan synthase 2 (HAS2) expression and hyaluronan secretion, presumably via putative CREB-binding sites adjacent to Smad-binding sites within the HAS2 promoter. Our findings identify the cAMP pathway as a potent but differential and promoter-specific regulator of TGF-beta-mediated effects involved in extracellular matrix homeostasis.
Department of Dermatology, Ludwig Boltzmann Institute for Cell Biology and Immunobiology of the Skin, 48149 Münster, Germany. schillm@uni-muenster.de
Meinhard
M
Schiller
Sylviane
S
Dennler
Ulf
U
Anderegg
Agatha
A
Kokot
Jan C
JC
Simon
Thomas A
TA
Luger
Alain
A
Mauviel
Markus
M
Böhm
D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D002465:Cell Movement / Q000187:drug effects; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D003094:Collagen / Q000096:biosynthesis; D055513:Connective Tissue Growth Factor / Q000096:biosynthesis; D000242:Cyclic AMP / Q000378:metabolism*; D020405:Dermis / Q000166:cytology; D050881:E1A-Associated p300 Protein / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D005109:Extracellular Matrix / Q000378:metabolism*; D016326:Extracellular Matrix Proteins / Q000235:genetics / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D006820:Hyaluronic Acid / Q000096:biosynthesis; D051379:Mice; D041681:NIH 3T3 Cells; D010766:Phosphorylation / Q000187:drug effects; D011480:Protease Inhibitors / Q000378:metabolism; D021381:Protein Transport / Q000187:drug effects; D020218:Response Elements / Q000235:genetics; D015398:Signal Transduction / Q000187:drug effects; D051785:Smad Proteins / Q000378:metabolism*; D016212:Transforming Growth Factor beta / Q000494:pharmacology*
C526317:CCN2 protein, human; D016326:Extracellular Matrix Proteins; D011480:Protease Inhibitors; D051785:Smad Proteins; D016212:Transforming Growth Factor beta; D055513:Connective Tissue Growth Factor; D006820:Hyaluronic Acid; D003094:Collagen; D000242:Cyclic AMP; D050881:E1A-Associated p300 Protein; C494360:EP300 protein, human; D000262:Adenylyl Cyclases