title: Altered signaling in the G1 phase deregulates chondrocyte growth in a mouse model with proteoglycan undersulfation.
abstract: In several skeletal dysplasias defects in extracellular matrix molecules affect not only the structural and mechanical properties of cartilage, but also the complex network of signaling pathways involved in cell proliferation and differentiation. Sulfated proteoglycans, besides playing an important structural role in cartilage, are crucial in modulating the transport, diffusion, and interactions of growth factors with their specific targets, taking part in the regulation of signaling pathways involved in skeletal development and growth. In this work, we investigated by real time PCR and Western blots of the microdissected growth plate and by immunohistochemistry the molecular basis of reduced chondrocyte proliferation in the growth plate of the dtd mouse, a chondrodysplastic model with defective chondroitin sulfate proteoglycan sulfation of articular and growth plate cartilage. We detected activation of the Wnt pathway, leading to an increase in the non-phosphorylated form of nuclear β-catenin and subsequent up-regulation of cyclin D1 expression in the G1 phase of the cell cycle. β-Catenin was further stabilized by up-regulation of Smad3 expression through TGF-β pathway synergistic activation. We demonstrate that notwithstanding cyclin D1 expression increase, cell cycle progression is compromised in the G1 phase due to reduced phosphorylation of the pocket protein p130 leading to inhibition of transcription factors of the E2F family which are crucial for cell cycle progression and DNA replication. These data, together with altered Indian hedgehox signaling detected previously, explain at the molecular level the reduced chondrocyte proliferation rate of the dtd growth plate leading to reduced skeletal growth.
authors.affiliation: Department of Molecular Medicine, Unit of Biochemistry, University of Pavia, Pavia, Italy.
authors.forename: Fabio
authors.initials: F
authors.lastname: De Leonardis
authors.forename: Luca
authors.initials: L
authors.lastname: Monti
authors.forename: Benedetta
authors.initials: B
authors.lastname: Gualeni
authors.forename: Ruggero
authors.initials: R
authors.lastname: Tenni
authors.forename: Antonella
authors.initials: A
authors.lastname: Forlino
authors.forename: Antonio
authors.initials: A
authors.lastname: Rossi
mesh_terms: D000818:Animals; D001846:Bone Development / Q000235:genetics*; D001847:Bone Diseases / Q000235:genetics; D001842:Bone and Bones / Q000378:metabolism / Q000473:pathology; D002358:Cartilage, Articular / Q000378:metabolism / Q000473:pathology; D002454:Cell Differentiation / Q000235:genetics; D049109:Cell Proliferation / Q000235:genetics; D019902:Chondrocytes / Q000378:metabolism*; D011508:Chondroitin Sulfate Proteoglycans / Q000378:metabolism; D019938:Cyclin D1 / Q000096:biosynthesis*; D050684:E2F Transcription Factors / Q000037:antagonists & inhibitors*; D005109:Extracellular Matrix / Q000473:pathology; D016193:G1 Phase / Q000235:genetics; D055879:Gene Knock-In Techniques; D006132:Growth Plate / Q000378:metabolism; D053823:Hedgehog Proteins / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D010766:Phosphorylation; D050717:Retinoblastoma-Like Protein p130 / Q000378:metabolism*; D015398:Signal Transduction / Q000235:genetics; D051900:Smad3 Protein / Q000096:biosynthesis; D016212:Transforming Growth Factor beta / Q000378:metabolism; D060449:Wnt Signaling Pathway; D051176:beta Catenin / Q000378:metabolism
chemical_list: D011508:Chondroitin Sulfate Proteoglycans; D050684:E2F Transcription Factors; D053823:Hedgehog Proteins; C493838:Rbl2 protein, mouse; D050717:Retinoblastoma-Like Protein p130; D051900:Smad3 Protein; C497791:Smad3 protein, mouse; D016212:Transforming Growth Factor beta; D051176:beta Catenin; C514061:ihh protein, mouse; D019938:Cyclin D1
keywords: CELL CYCLE; GROWTH PLATE; PROTEOGLYCAN SULFATION; SKELETAL DYSPLASIA; WNT PATHWAY