Identification of endoglin in rat hepatic stellate cells: new insights into transforming growth factor beta receptor signaling.
Transforming growth factor beta (TGF-beta) signaling is mediated by the cell surface TGF-beta type I (ALK5), type II, and the accessory type III receptors endoglin and betaglycan. Hepatic stellate cells (HSC), the most profibrogenic cell type in the liver, express ALK5, TbetaRII, and betaglycan. To monitor the expression of betaglycan in HSC, we used the commercially available antibody sc-6199 in Western blot analysis. This antibody, raised against a peptide mapping at the carboxyl terminus of the human betaglycan, is claimed to be specific for betaglycan, although it is known that the C-terminal domain is highly conserved in type III receptors. Proteins recognized in HSC by sc-6199 did not match the characteristic migration pattern of betaglycan. Moreover, the determined molecular weight (M(r) 160) and the observed reductant sensitivity after treatment with dithiothreitol resemble those of a closely related type III receptor, endoglin (CD105). Endoglin, a disulfide-linked homodimer, is an accessory component of the TGF-beta receptor complex and mainly expressed on endothelial cells. The presence of endoglin in HSC of rat liver was confirmed by molecular cloning of the endoglin cDNA and immunocytochemistry. The reactivity of sc-6199 with both auxiliary TGF-beta receptors (betaglycan and endoglin) from rats was demonstrated by Western blot and immunocytochemical analysis of cells heterologously expressing these proteins. Furthermore, Northern and Western blotting revealed that both betaglycan and endoglin genes are differentially regulated in HSC and in transdifferentiated myofibroblasts (MFB). By surface labeling and immunoprecipitation experiments, we show that endoglin is found in significant amounts exposed at the plasma membrane of HSC and MFB, which is a pivotal prerequisite for binding of and signaling in response to TGF-beta. In conclusion, we hypothesize that TGF-beta signals in HSC and MFB are tuned by two different interconnected signaling pathways, as it was previously demonstrated for endothelial cells.
Meurer
Steffen K
SK
Institute of Clinical Chemistry and Pathobiochemistry, RWTH-University Hospital, D-52074 Aachen, Germany.
Tihaa
Lidia
L
Lahme
Birgit
B
Gressner
Axel M
AM
Weiskirchen
Ralf
R
D000595:Amino Acid Sequence; D000818:Animals; D015703:Antigens, CD; D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D019556:COS Cells; D002454:Cell Differentiation; D018076:DNA, Complementary / Q000378:metabolism; D019281:Dimerization; D000071063:Endoglin; D005347:Fibroblasts / Q000166:cytology; D005786:Gene Expression Regulation; D007150:Immunohistochemistry; D047468:Immunoprecipitation; D008099:Liver / Q000166:cytology*; D008297:Male; D008957:Models, Genetic; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D017434:Protein Structure, Tertiary; D011509:Proteoglycans / Q000737:chemistry; D051381:Rats; D017207:Rats, Sprague-Dawley; D011956:Receptors, Cell Surface; D018125:Receptors, Transforming Growth Factor beta / Q000737:chemistry / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D013997:Time Factors; D014162:Transfection; D019010:Vascular Cell Adhesion Molecule-1 / Q000737:chemistry* / Q000378:metabolism
D015703:Antigens, CD; D018076:DNA, Complementary; C064206:ENG protein, human; D000071063:Endoglin; D011509:Proteoglycans; D011956:Receptors, Cell Surface; D018125:Receptors, Transforming Growth Factor beta; D019010:Vascular Cell Adhesion Molecule-1; C071736:betaglycan