Miller Stephanie A, Brown Anissa J, Farach-Carson Mary C, Kirn-Safran Catherine B
Department of Biological Sciences, University of Delaware, 310 Wolf Hall, Newark, DE 19716, USA.
Differentiation. 2003 Aug;71(6):322-36. doi: 10.1046/j.1432-0436.2003.7106002.x.
HIP is a heparin/heparan sulfate (Hp/HS) binding protein identical to ribosomal protein L29 that displays diverse biological functions. There is strong evidence that abnormal expression and quantitative deficiencies of essential molecules such as extracellular matrix (ECM) proteins, transcription factors, and ribosomal proteins can seriously impair embryonic development. As observed for HS-bearing molecules, high levels of HIP/RPL29 are found in proliferating chondrocytic precursors and chondrocytes of developing growth plate. Here, we demonstrate both in vitro and in developing mouse embryos that HIP/RPL29 is down-regulated in terminally differentiated chondrocytes corresponding to the late hypertrophic zone of the growth plate. Because cartilage serves as a template for endochondral bone formation, we hypothesize that the presence of HIP/RPL29 during early chondrogenesis is essential for normal skeletal growth and patterning. In particular, we believe that HIP/RPL29 expression is required to maintain proliferation of chondrocytes and avoid skeletal shortening. Increasing evidence suggests that multifunctional ribosomal proteins of eukaryotic cells are important regulators of cell growth and differentiation, not simply structural parts of translational machinery. To investigate the role of HIP/RPL29 normal expression during cartilage formation, we designed a ribozyme-mediated knock-down approach to partially down-regulate HIP/RPL29 expression in the multipotent mouse embryonic skin fibroblast cell line C3H/10T (1/2). This technology permitted us to avoid the insufficient expression associated with more severe consequences, such as lethality, and provided advantages similar to those obtained with mutations generating hypomorphic phenotypes. Our results show that partial reduction of HIP/RPL29 levels accelerates differentiation of C3H/10T(1/2) into cartilage-like cells. In conclusion, our data indicate that HIP/RPL29 constitutes an important novel regulator of chondrocytic growth and differentiation.
HIP是一种与核糖体蛋白L29相同的肝素/硫酸乙酰肝素（Hp/HS）结合蛋白，具有多种生物学功能。有充分证据表明，细胞外基质（ECM）蛋白、转录因子和核糖体蛋白等必需分子的异常表达和定量缺陷会严重损害胚胎发育。正如在含HS分子中观察到的那样，在发育中的生长板的增殖软骨细胞前体和软骨细胞中发现了高水平的HIP/RPL29。在这里，我们在体外和发育中的小鼠胚胎中均证明，在与生长板晚期肥大区相对应的终末分化软骨细胞中，HIP/RPL29表达下调。由于软骨是软骨内骨形成的模板，我们推测早期软骨形成过程中HIP/RPL29的存在对于正常骨骼生长和模式形成至关重要。特别是，我们认为需要HIP/RPL29表达来维持软骨细胞的增殖并避免骨骼缩短。越来越多的证据表明，真核细胞的多功能核糖体蛋白是细胞生长和分化的重要调节因子，而不仅仅是翻译机器的结构部分。为了研究HIP/RPL29正常表达在软骨形成过程中的作用，我们设计了一种核酶介导的敲低方法，以部分下调多能小鼠胚胎皮肤成纤维细胞系C3H/10T（1/2）中HIP/RPL29的表达。这项技术使我们能够避免与更严重后果（如致死性）相关的表达不足，并提供了与产生亚等位基因表型的突变所获得的优势相似的优势。我们的结果表明，HIP/RPL29水平的部分降低会加速C3H/10T（1/2）向软骨样细胞的分化。总之，我们的数据表明HIP/RPL29构成了软骨细胞生长和分化的重要新型调节因子。
