title: Notch signaling indirectly promotes chondrocyte hypertrophy via regulation of BMP signaling and cell cycle arrest.
abstract: Cell cycle regulation is critical for chondrocyte differentiation and hypertrophy. Recently we identified the Notch signaling pathway as an important regulator of chondrocyte proliferation and differentiation during mouse cartilage development. To investigate the underlying mechanisms, we assessed the role for Notch signaling regulation of the cell cycle during chondrocyte differentiation. Real-time RT-PCR data showed that over-expression of the Notch Intracellular Domain (NICD) significantly induced the expression of p57, a cell cycle inhibitor, in chondrocytes. Flow cytometric analyses further confirmed that over-expression of NICD in chondrocytes enhances the G0/G1 cell cycle transition and cell cycle arrest. In contrast, treatment of chondrocytes with the Notch inhibitor, DAPT, decreased both endogenous and BMP2-induced SMAD 1/5/8 phosphorylation and knockdown of SMAD 1/5/8 impaired NICD-induced chondrocyte differentiation and p57 expression. Co-immunoprecipitation using p-SMAD 1/5/8 and NICD antibodies further showed a strong interaction of these proteins during chondrocyte maturation. Finally, RT-PCR and Western blot results revealed a significant reduction in the expression of the SMAD-related phosphatase, PPM1A, following NICD over-expression. Taken together, our results demonstrate that Notch signaling induces cell cycle arrest and thereby initiates chondrocyte hypertrophy via BMP/SMAD-mediated up-regulation of p57.
authors.affiliation: Department of Orthopedic Surgery, Anhui Provincial Hospital, 17 Lujiang Rd, Hefei, China.|Department of Orthopedic Surgery, Louisiana State University Health Sciences Center, Shreveport, Louisiana, USA.
authors.forename: Xifu
authors.initials: X
authors.lastname: Shang
authors.affiliation: Shanghai Key Laboratory of Orthopedic Implant, Department of Orthopaedic Surgery, Shanghai Ninth People's Hospital Affiliated Shanghai Jiao Tong University School of Medicine, Shanghai, China.
authors.forename: Jinwu
authors.initials: J
authors.lastname: Wang
authors.affiliation: Department of Orthopedic Surgery, Anhui Provincial Hospital, 17 Lujiang Rd, Hefei, China.
authors.forename: Zhengliang
authors.initials: Z
authors.lastname: Luo
authors.affiliation: Institute of Spine, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai, China.
authors.forename: Yongjun
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Department of Orthopedic Surgery, Louisiana State University Health Sciences Center, Shreveport, Louisiana, USA.
authors.forename: Massimo M
authors.initials: MM
authors.lastname: Morandi
authors.affiliation: Department of Orthopedic Surgery, Louisiana State University Health Sciences Center, Shreveport, Louisiana, USA.
authors.forename: John V
authors.initials: JV
authors.lastname: Marymont
authors.affiliation: Departments of Orthopedic Surgery and Cell Biology, Duke Orthopedic Cellular, Developmental, and Genome Laboratories, Duke University School of Medicine, Durham, North Carolina, USA.
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Hilton
authors.affiliation: Department of Orthopedic Surgery, Louisiana State University Health Sciences Center, Shreveport, Louisiana, USA.
authors.forename: Yufeng
authors.initials: Y
authors.lastname: Dong
mesh_terms: D000818:Animals; D055396:Bone Morphogenetic Protein 2 / Q000235:genetics* / Q000378:metabolism; D059447:Cell Cycle Checkpoints / Q000235:genetics*; D002454:Cell Differentiation / Q000235:genetics; D049108:Cell Enlargement; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000235:genetics; D019902:Chondrocytes / Q000378:metabolism* / Q000473:pathology; D050761:Cyclin-Dependent Kinase Inhibitor p57 / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation; D051379:Mice; D010766:Phosphorylation; D034622:RNA Interference; D051880:Receptors, Notch / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction / Q000235:genetics; D051896:Smad Proteins, Receptor-Regulated / Q000235:genetics / Q000378:metabolism
chemical_list: C525719:Bmp2 protein, mouse; D055396:Bone Morphogenetic Protein 2; C493965:Cdkn1c protein, mouse; D050761:Cyclin-Dependent Kinase Inhibitor p57; D051880:Receptors, Notch; D051896:Smad Proteins, Receptor-Regulated