The use of genetic models to define the role of beta-catenin in post-natal growth

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DESCRIPTION (provided by applicant): ?-catenin has emerged as a critical regulator of endochondral bone formation. However, the ability to study the role of ?-catenin in growth and development is severely limited by the embryonic or early lethality that occurs in all cartilage-related gene deletions that result in loss or gain of ?-catenin function. This proposal defines the role of ?-catenin in post-natal growth and development and uses an innovative approach that permits us to temporally control tissue specific conditional gene deletion in mice. Preliminary data establish that delivery of tamoxifen to Col2a1-Cre+/-ERT2 transgenic mice results in tissue specific gene recombination. Aim 1 characterizes the temporal expression of Cre-recombinase in murine cartilage and assesses the role of ?-catenin gain of function on post-natal growth and development. Aim 1A consists of a focused set of experiments that complete a characterization of the pattern of gene recombination following delivery of tamoxifen and optimizes its administration using Col2a1-Cre+/-ERT2; ROSA26R +/- mice. Aim 1B examines the phenotype of ?-catenin+/fiox(exon3); Col2a1-Cre+/-ERT2 mice in which post-natal conditional gene deletion at 2 weeks results in expression of a constitutively active ?-catenin in cartilage. Aim 2 characterizes the role of ?-catenin loss of function on post-natal growth and development and uses two complementary models. ICAT is an intracellular protein that competitively binds ?-catenin and inhibits signaling. Preliminary data show delayed chondrocyte maturation and runting in Col2a1-ICAT transgenic mice, which have inhibition of ?-catenin signaling throughout development and during post-natal growth. Aim 2A characterizes the phenotype of Col2a1-ICAT transgenic mice. Aim 2B characterizes the phenotype of ?-cateninfloxKO/ floxKO; Col2a1-Cre+/-ERT2 mice in which post-natal conditional gene deletion at 2 weeks results in loss of ?-catenin expression in cartilage. Aim 3 uses an in vitro approach and defines mechanisms involved in the interdependence of ?-catenin and BMP signaling on VEGF gene expression during terminal chondrocyte maturation. Preliminary data show that induction of VEGF requires both BMP and ?-catenin signaling. Aim 3A examines regulation of BMP-2, 4, and 6 expressions in chondrocytes by ?-catenin. Aim 3B defines the cooperative induction of the VEGF gene by BMP-2 and ?-catenin. Aim 3C defines critical Smad-?-catenin interactions on the VEGF-A promoter. Altogether, these experiments will establish the concept that regulation of key maturation associated genes by ?-catenin requires the presence of BMP co-signals that act to enhance the rate of chondrocyte differentiation. The findings will establish ?-catenin and its cooperative effects with BMPs as an essential signal for chondrocyte hypertrophy, completion of endochondral bone formation, and regulation of VEGF expression. The proposed studies will provide critical new insights regarding cartilage diseases including chondrodysplasia, bone repair, and osteoarthritis.