The function of R-spondins during mouse limb development and chondrogenesis

一组称为WNTs、刺猬、BMPs和FGFs的细胞信号控制着大部分胚胎发育。这些分泌的蛋白质以精确的空间和时间序列相互作用，形成胚胎的模式。肢体(胳膊和腿)的形成是脊椎动物最具特点的发育途径之一。我们使用小鼠的肢体作为一个系统来理解细胞信号如何控制形态发生(形态的创造)和细胞分化。这些信号系统的活动被精确调节的机制尚不清楚。近年来，R-Respondin蛋白(RSPO)已被确定为WNT途径的重要组成部分。RSPO是在肢体发育过程中修改这些关键通路的关键候选者。编码RSPO的四个基因(Rsp1-4)已被证明具有从性别决定到胎盘、肺和肢体发育的重要发育功能。随着必要的小鼠突变体的出现，肢体形态发生很可能成为第一个全面描述RSPO基因功能的发育系统。这项建议通过围绕肢体发育的连续阶段组织两个目标来追求这一目标：1)肢体芽的早期生长和模式；2)软骨分化和成熟的后期过程，建立肢体骨架的软骨模板。在目标1中，我们确定了Rsp4在肢体发育过程中的功能以及它与Rsp2和Rsp3的遗传相互作用。通过结合Rspo4和Rspo2/3突变，我们将对这些基因在肢体发育过程中的功能有更全面的了解。在目标2中，我们使用一种称为微团细胞培养的体外系统来确定R-Spinins在肢体软骨发育过程中的功能。在它们参与肢体早期发育后，我们假设R-Spinins在软骨产生细胞--软骨细胞的分化过程中发挥作用。综上所述，这些目标将揭示R-Spinins如何帮助控制WNT信号通路，以便在发育过程中以准确的方式部署它们。

A small set of cellular signals called WNTs, hedgehogs, BMPs, and FGFs control much of embryonic development. These secreted proteins interact in a precise spatial and temporal sequence to pattern the embryo. The formation of the limbs (arms and legs) is among the best-characterized developmental pathways in vertebrates. We use the mouse limb as a system to understand how cellular signals control morphogenesis (the creation of form) and cellular differentiation. The mechanisms by which the activities of these signaling systems are precisely modulated is unknown. Recently, the R-spondin proteins (RSPOs) have been identified as important components of the WNT pathways. RSPOs are key candidates for modifying these critical pathways during limb development. The four genes that code for RSPOs (Rspo1-4) have been shown to have important developmental functions ranging from sex determination to placental, lung and limb development. As the necessary mouse mutants are becoming available, limb morphogenesis will likely be the first developmental system where a comprehensive description of Rspo gene function may be obtainable. This proposal pursues this goal by organizing two objectives around successive stages of limb development: 1) The early outgrowth and patterning of the limb bud and 2) the later process of chondrogenic differentiation and maturation that establishes the cartilage template of the limb skeleton. In Objective 1 we determine the function of Rspo4 during limb development and its genetic interactions with Rspo2 and Rspo3. By combining the Rspo4 and Rspo2/3 mutations we will obtain a more complete understanding of the function of these genes during limb development. In Objective 2 we determine the function of R-spondins during limb cartilage development, using an in vitro system called micromass cell culture. After they have participated in early limb development, we hypothesize that the R-spondins have a function during the differentiation of the cartilage-producing cells, the chondrocytes. Taken together these aims will reveal how R-spondins help control the WNT signaling pathways so that they are deployed in a precise manner during development.