Abstract
Gene-trap mutagenesis is based on the notion that the random insertion of a trapping vector may disturb the function of inserted genes. Here, we applied this method to murine mesenchymal ATDC5 cells, which differentiate into mature chondrocytes in the presence of insulin. As the trap vector we used pPT1-geo, which lacks its own promoter and enhancer, but contains a lacZ-neo fusion gene as a reporter and selection marker driven by the promoter of the trapped gene. After pPT1-geo was introduced into ATDC5 cells by electroporation, the neomycin-resistant clones were screened for beta-galactosidase activity. The selected clones were cultured in differentiation medium to evaluate the chondrogenic phenotype. The clones no. 6-30 and 6-175, which exhibited impaired and accelerated mineralization, respectively, were subjected to further analysis. In clone no. 6-30 in which the gene coding for the p85alpha subunit of phosphoinositide 3-kinase (PI3K) was trapped, the expression of marker genes of early chondrocytes including collagen type II, aggrecan, and PTH/PTHrP receptor was delayed. The insulin-induced stimulation of growth was reduced in clone no. 6-30 compared with the parental ATDC5 cells. Moreover, treatment of parental ATDC5 cel...Continue Reading
References
May 1, 1990·Cell Differentiation and Development : the Official Journal of the International Society of Developmental Biologists·T AtsumiY Ikawa
Aug 1, 1995·Molecular and Cellular Biology·S PonsM F White
Feb 1, 1994·Genes & Development·A C KaraplisR C Mulligan
Apr 1, 1996·The Journal of Cell Biology·C ShukunamiY Hiraki
Mar 8, 1996·The Journal of Biological Chemistry·K InukaiT Asano
Aug 2, 1996·Science·A VortkampC J Tabin
Aug 2, 1996·Science·B LanskeH M Kronenberg
Jan 1, 1996·Journal of Bone and Mineral Research : the Official Journal of the American Society for Bone and Mineral Research·H AkiyamaT Nakamura
Oct 1, 1996·Genomics·D A FrumanC L Carpenter
Mar 21, 1997·The Journal of Biological Chemistry·K InukaiT Asano
Jun 12, 1997·Nature·A Toker, L C Cantley
Aug 1, 1997·Journal of Bone and Mineral Research : the Official Journal of the American Society for Bone and Mineral Research·C ShukunamiY Hiraki
Jan 15, 1999·Science·H SuzukiS Koyasu
Jan 15, 1999·Science·D A FrumanL C Cantley
Feb 13, 1999·Nature Genetics·Y TerauchiT Kadowaki
Aug 31, 1999·Genes & Development·B St-JacquesA P McMahon
Jul 11, 2000·Nature Biotechnology·G MainguyM Volovitch
Jul 19, 2000·FEBS Letters·P BorgattiL M Neri
Oct 5, 2001·Nature Reviews. Genetics·W L StanfordS P Cordes
Nov 8, 2001·Current Opinion in Cell Biology·B de CrombruggheK Nakashima
Apr 9, 2002·Genes & Development·Zhonghao LiuDavid M Ornitz
Apr 9, 2002·Genes & Development·Norihiko OhbayashiShinji Takada
Jun 27, 2002·The Journal of Biological Chemistry·Nandini Ghosh-ChoudhuryGoutam Ghosh Choudhury
Oct 4, 2002·Developmental Cell·Eleonora MininaAndrea Vortkamp
Jun 5, 2003·Genes & Development·Xiao-Ding PengNissim Hay
Citations
Mar 15, 2012·The Journal of Biological Chemistry·Takao KoshimizuToshimi Michigami
Aug 17, 2010·Bone·Masaaki KimataKeiichi Ozono
Mar 15, 2008·Biochemical and Biophysical Research Communications·Hiroaki WatanabeB Mark Evers
Oct 2, 2007·Bone·Takayuki UchihashiToshimi Michigami
Dec 4, 2009·The Anatomical Record : Advances in Integrative Anatomy and Evolutionary Biology·Erik Kern HarringtonKathy K H Svoboda
Feb 27, 2010·Arthritis and Rheumatism·Atsushi FukaiHiroshi Kawaguchi
Aug 28, 2009·Journal of Cellular Physiology·Chin-Chang ChengMakarand V Risbud
Aug 18, 2020·Biomedical Research·Wataru AriyoshiTatsuji Nishihara
Apr 6, 2011·Journal of Cellular Biochemistry·Byung-Gyu KimJe-Yoel Cho