ROLES OF SOX C GENES IN SKELETOGENESIS

描述(申请人提供)：该项目将揭示Sox4、Sox11和Sox12在体内骨骼细胞中的作用。这三个高度相关的蛋白形成了Sry相关HMG盒(SOX)转录因子的C亚家族。它们的基因在整个小鼠胚胎的骨源性间充质细胞中共表达，并在这些细胞经历软骨细胞和成骨细胞分化时保持差异表达。虽然Sox12在体内的作用尚不清楚，但缺乏Sox4和/或Sox11的小鼠已经开始揭示这两个基因在骨骼形成的多个步骤中的重要作用。Sox C蛋白的分子作用在很大程度上仍不清楚，但初步研究表明，它们可能与DNA结合并激活转录，但效率不同。这些数据提出了一种假设，即SOX C蛋白可能在控制骨骼细胞命运和分化方面具有冗余、互补或独特的分子作用。为了验证这一假设，目标1是进一步研究Sox C蛋白在小鼠胚胎骨骼形成中的作用。携带Sox11和Sox12条件零等位基因的小鼠将使用与生成Sox4条件零等位基因相同的策略来生成。成熟的Cre转基因基因将被用来在整个胚胎中或在骨骼形成过程中的特定步骤中特异性地使基因失活。骨骼缺陷将在形态、细胞和分子水平上进行表征，并将进行原代细胞培养实验，以补充体内研究。目的2研究SOX C蛋白在骨骼形成细胞中的分子作用。三种Sox C蛋白的DNA结合和反式激活特性将使用标准的体外和细胞培养方法进一步研究。潜在的靶基因将通过基因表达阵列筛选来确定。SOX C蛋白对靶基因调控序列的作用将通过体外和体内的DNA结合和反式激活分析来研究。预计这项研究将确定SOX C蛋白是特定细胞谱系中从多能前体阶段到后期成熟的骨骼形成细胞的基本决定因素，从而为了解正常骨骼形成以及骨骼的遗传和获得性疾病所涉及的分子机制提供重要的见解。

DESCRIPTION (provided by applicant): This project will uncover the roles of Sox4, Sox11, and Sox12 in skeletogenic cells in vivo. These three highly related proteins form the subfamily C of Sry-related HMG box (Sox) transcription factors. Their genes are co-expressed in skeletogenic mesenchymal cells throughout the mouse embryo, and remain differentially expressed when these cells undergo chondrocyte and osteoblast differentiation. While the roles of Sox12 in vivo remain unknown, mice lacking Sox4 and/or Sox11 have started to reveal important roles for these two genes in multiple steps of skeletogenesis. The molecular roles of Sox C proteins remain largely unknown, but preliminary studies have suggested that they may bind DNA and activate transcription similarly but with different efficiencies. These data raise the hypothesis that Sox C proteins may have redundant, complementary, or distinct molecular roles in controlling skeletogenic cell fate and differentiation. To test this hypothesis, Aim 1 is to further study the roles of the Sox C proteins in mouse embryo skeletogenesis. Mice carrying Sox11 and Sox12 conditional null alleles will be generated using the same strategy that was used to generate a Sox4 conditional null allele. Well-established Cre transgenes will be used to specifically inactivate the genes in the entire embryo or at specific steps during skeletogenesis. Skeletal defects will be characterized at the morphological, cellular and molecular levels, and primary cell culture experiments will be carried out to complement in vivo studies. Aim 2 is to characterize the molecular roles of Sox C proteins in skeletogenic cells. The DNA-binding and transactivation properties of the three Sox C proteins will be further studied using standard in vitro and cell culture assays. Potential target genes will be identified by gene expression array screening. The action of Sox C proteins on target gene regulatory sequences will be studied using DNA-binding and transactivation assays in vitro and in vivo. It is anticipated that this study will identify Sox C proteins as essential determinants of skeletogenic cells from pluripotent precursor stages until late maturation in specific cell lineages, and will thereby provide significant insights into molecular mechanisms involved in normal skeletogenesis and in genetic and acquired diseases of the skeleton.