Hepatocyte growth factor suppresses profibrogenic signal transduction via nuclear export of Smad3 with galectin-7.
OBJECTIVE
Hepatocyte growth factor (HGF) and transforming growth factor-beta (TGF-beta) regulate diversified cellular functions and often act antagonistically against each other. For example, TGF-beta is the most potent factor accelerating liver fibrosis, whereas HGF treatment prevents its progression. Here, we propose a novel molecular mechanism by which HGF counter represses TGF-beta-stimulated profibrogenic signal transduction.


METHODS
Effects of HGF on TGF-beta-responsive gene transcription of type I collagen, the major matrix component of fibrotic liver, were examined by using cultured hepatic stellate cells (HSC) and transgenic mice harboring alpha2(I) collagen gene (COL1A2) promoter. Expression and subcellular localization of Smad3 were determined by Western blot analyses and immunofluorescence staining, respectively. A mass spectrometric analysis was employed to identify immunoprecipitated proteins with antiphospho-Smad2/3 antibodies.


RESULTS
Over expression of HGF inhibited COL1A2 transcription in cultured HSC and suppressed activation of COL1A2 promoter in liver tissue induced by carbon tetrachloride administration. A mass spectrometric analysis identified galectin-7 as one of the immunoprecipitated proteins with antiphospho-Smad2/3 antibodies following HGF treatment. HGF accelerated nuclear export of Smad3 by enhancing its interaction with galectin-7. Transfection of cells with galectin-7 small interfering RNA inhibited nuclear export of Smad3 and abolished suppressive effect of HGF on expression of TGF-beta-responsive genes such as COL1A2 and plasminogen activator inhibitor-1. On the other hand, over expression of galectin-7 suppressed TGF-beta-stimulated expression of those target genes.


CONCLUSIONS
These results reveal a novel function of intracellular galectin-7 as a transcriptional regulator via its interaction with Smad3 and provide a molecular basis for the antifibrotic effect of HGF.
Inagaki
Yutaka
Y
Liver Fibrosis Research Unit, Department of Community Health, Tokai University School of Medicine, Isehara, Japan. yutakai@is.icc.u-tokai.ac.jp
Higashi
Kiyoshi
K
Kushida
Miwa
M
Hong
Yun Yu
YY
Nakao
Sachie
S
Higashiyama
Reiichi
R
Moro
Tadashi
T
Itoh
Johbu
J
Mikami
Toshiyuki
T
Kimura
Toru
T
Shiota
Goshi
G
Kuwabara
Ichiro
I
Okazaki
Isao
I
D000818:Animals; D000906:Antibodies / Q000032:analysis; D015153:Blotting, Western; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors; D002478:Cells, Cultured; D003094:Collagen / Q000096:biosynthesis / Q000235:genetics; D024042:Collagen Type I; D018450:Disease Progression; D004789:Enzyme Activation / Q000187:drug effects; D005419:Flavonoids / Q000494:pharmacology; D005455:Fluorescent Antibody Technique; D037161:Galectins / Q000096:biosynthesis / Q000235:genetics*; D015870:Gene Expression; D017228:Hepatocyte Growth Factor / Q000096:biosynthesis / Q000235:genetics / Q000627:therapeutic use*; D047468:Immunoprecipitation; D008106:Liver Cirrhosis, Experimental / Q000235:genetics / Q000378:metabolism / Q000517:prevention & control*; D013058:Mass Spectrometry; D051379:Mice; D008822:Mice, Transgenic; D018613:Microscopy, Confocal; D048052:Mitogen-Activated Protein Kinase 3 / Q000187:drug effects / Q000378:metabolism; D017395:Plasminogen Activator Inhibitor 1 / Q000096:biosynthesis / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000187:drug effects; D012313:RNA / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects*; D051900:Smad3 Protein / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology; D014158:Transcription, Genetic / Q000187:drug effects
D000906:Antibodies; D024042:Collagen Type I; D005419:Flavonoids; D037161:Galectins; C490487:Lgals7 protein, mouse; D017395:Plasminogen Activator Inhibitor 1; D051900:Smad3 Protein; C497791:Smad3 protein, mouse; D012313:RNA; D017228:Hepatocyte Growth Factor; D003094:Collagen; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D048052:Mitogen-Activated Protein Kinase 3; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one