PTHrP expression in chick sternal chondrocytes is regulated by TGF-beta through Smad-mediated signaling.
PTHrP regulates the rate of chondrocyte differentiation during endochondral bone formation. The expression of PTHrP and its regulation by TGF-beta, BMP-2, and PTHrP was examined in upper sternal chondrocytes following 1, 3, and 5 days of continuous treatment. While TGF-beta stimulated the expression of PTHrP (5-fold), PTHrP caused a slight inhibition, and BMP-2 markedly inhibited PTHrP mRNA expression. The effect of these factors on PTHrP expression was not simply related to the maturational state of the cells, since BMP-2 increased, while both PTHrP and TGF-beta decreased the expression of type X collagen. TGF-beta isoforms 1, 2, and 3 all stimulated PTHrP expression. Signaling events involved in the induction of PTHrP by TGF-beta were further evaluated in a PTHrP-promoter CAT construct. The effect of TGF-beta, BMP-2, and PTHrP on the PTHrP-promoter paralleled their effects on mRNA expression, with TGF-beta significantly increasing CAT activity, BMP-2 decreasing CAT activity, and PTHrP having a minimal effect. Co-transfection of the TGF-beta signaling molecule, Smad 3, mimicked the effect of TGF-beta (induction of PTHrP promoter), while dominant negative Smad 3 inhibited the induction of the PTHrP promoter by TGF-beta. Furthermore, infection with a Smad 3-expressing retrovirus mimicked the effects of exogenously added TGF-beta, and induced PTHrP mRNA expression in the infected chondrocyte culture. In contrast, a dominant negative Smad 3 completely inhibited PTHrP promoter stimulation by TGF-beta, but only partially blocked the effect of TGF-beta on PTHrP mRNA synthesis. These findings demonstrate that PTHrP is expressed in chondrocytes undergoing endochondral ossification, and show regulation, at least in part, by TGF-beta through Smad mediated signaling events.
Pateder
D B
DB
Center for Musculoskeletal Research, University of Rochester School of Medicine and Dentistry, Rochester, New York 14642, USA.
Ferguson
C M
CM
Ionescu
A M
AM
Schwarz
E M
EM
Rosier
R N
RN
Puzas
J E
JE
O'Keefe
R J
RJ
D000818:Animals; D002454:Cell Differentiation / Q000502:physiology; D002478:Cells, Cultured; D002642:Chick Embryo; D002645:Chickens; D019902:Chondrocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects / Q000502:physiology; D044162:Parathyroid Hormone-Related Protein; D011401:Promoter Regions, Genetic / Q000187:drug effects; D014176:Protein Biosynthesis; D011506:Proteins / Q000235:genetics / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D051900:Smad3 Protein; D013249:Sternum; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D015533:Transcriptional Activation / Q000187:drug effects; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000378:metabolism* / Q000494:pharmacology
D004268:DNA-Binding Proteins; D044162:Parathyroid Hormone-Related Protein; D011506:Proteins; D012333:RNA, Messenger; D051900:Smad3 Protein; D015534:Trans-Activators; D016212:Transforming Growth Factor beta