title: Vinculin functions as regulator of chondrogenesis.
abstract: To identify the genes involved in chondrocytic differentiation, we applied gene trap mutagenesis to a murine mesenchymal chondrogenic cell line ATDC5 and isolated a clone in which the gene encoding vinculin was trapped. The trapped allele was assumed to express a fusion protein containing a truncated vinculin lacking the tail domain and the geo product derived from the trap vector. The truncated vinculin was suggested to exert a dominant negative effect. Impaired functioning of vinculin caused by gene trapping in ATDC5 cells or knockdown in primary chondrocytes resulted in the reduced expression of chondrocyte-specific genes, including Col2a1, aggrecan, and Col10a1. The expression of Runx2 also was suppressed by the dysfunctional vinculin. On the other hand, the expression of Sox9, encoding a key transcription factor for chondrogenesis, was retained. Knockdown of vinculin in metatarsal organ cultures impaired the growth of the explants and reduced the expression of Col2a1 and aggrecan. Gene trapping or knockdown of vinculin decreased the phosphorylation of ERK1/2 but increased that of Src homology 2 domain-containing tyrosine phosphatase 2 (SHP2) and Akt during chondrocytic differentiation, suggesting a disturbance of signaling by insulin-like growth factor I (IGF-I). Knockdown of vinculin in the metatarsal organ culture abrogated the IGF-I-induced growth and inhibited the up-regulation of Col2a1 and aggrecan expression by IGF-I. Loss of vinculin function in differentiating chondrocytes impaired the activation of the p38 MAPK pathway also, suggesting its involvement in the regulation of chondrogenesis by vinculin. Our results indicate a tissue-specific function of vinculin in cartilage whereby it controls chondrocytic differentiation.
authors.affiliation: Department of Bone and Mineral Research, Osaka Medical Center and Research Institute for Maternal and Child Health, 840 Murodo-cho, Izumi, Osaka 594-1101, Japan.
authors.forename: Takao
authors.initials: T
authors.lastname: Koshimizu
authors.forename: Masanobu
authors.initials: M
authors.lastname: Kawai
authors.forename: Hiroki
authors.initials: H
authors.lastname: Kondou
authors.forename: Kanako
authors.initials: K
authors.lastname: Tachikawa
authors.forename: Norio
authors.initials: N
authors.lastname: Sakai
authors.forename: Keiichi
authors.initials: K
authors.lastname: Ozono
authors.forename: Toshimi
authors.initials: T
authors.lastname: Michigami
mesh_terms: D053674:Aggrecans / Q000235:genetics / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western; D019556:COS Cells; D002454:Cell Differentiation / Q000235:genetics / Q000502:physiology*; D002460:Cell Line; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D019902:Chondrocytes / Q000166:cytology / Q000378:metabolism*; D020219:Chondrogenesis; D002999:Clone Cells; D024043:Collagen Type II / Q000235:genetics / Q000378:metabolism; D024121:Collagen Type X / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D055785:Gene Knockdown Techniques; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology; D008682:Metatarsal Bones / Q000254:growth & development / Q000378:metabolism; D051379:Mice; D009154:Mutation; D009924:Organ Culture Techniques; D034622:RNA Interference; D020133:Reverse Transcriptase Polymerase Chain Reaction; D055758:SOX9 Transcription Factor / Q000235:genetics / Q000378:metabolism; D016596:Vinculin / Q000235:genetics / Q000378:metabolism / Q000502:physiology*
chemical_list: D053674:Aggrecans; C570620:Col10a1 protein, mouse; C447413:Col2a1 protein, mouse; D024043:Collagen Type II; D024121:Collagen Type X; D055758:SOX9 Transcription Factor; C527933:Sox9 protein, mouse; D016596:Vinculin; D007334:Insulin-Like Growth Factor I