Role of BMP and FGF signaling during limb development

发育中的一个基本问题是确定细胞如何决定保持多能性或分化。这个问题存在于胚胎中如何产生模式的核心，反之，当这些过程被破坏时，如在肿瘤形成中，模式如何被破坏。发育中的肢体是解决这些问题的理想系统，因为由于遗传学和实验胚胎学的研究，存在着关于肢体发育的丰富数据。此前，我们已经通过研究缺乏编码成纤维细胞生长因子配体的基因的小鼠，展示了成纤维细胞生长因子信号在肢体发育中的重要作用(Lewis等人)。2000自然遗传学28：167，Sun等人2000自然遗传学25：6)。然而，由成纤维细胞生长因子基因冗余引起的复杂性使我们考虑通过检测成纤维细胞生长因子受体突变来解决这一问题，从而得出成纤维细胞生长因子信号控制肢体大小的见解(Verheyden等人，2005Development 132：4235)。通过骨形态发生蛋白(BMPs)调控的信号通路被认为在肢体生长的各个方面发挥作用：三个轴的早期图案、细胞程序性死亡和骨形成。摆在我们面前的任务是了解BMP和成纤维细胞生长因子信号通路在肢体发育过程中如何相互作用。BMP被认为在其中发挥直接作用的一个过程是，作为发生在间充质指间细胞中的正常程序性细胞死亡的效应器，从而去除它们，并在出生时没有鳍状肢体的动物中塑造最终的手指图案。然而，我们在最近的出版物(Pajni-Underwood S.et al 2007 Development 134：2359)中挑战了这一范式。通过同时失活BMP受体基因BMPR1A以及特定于肢芽外胚层的Fgf8和Fgf4，我们已经为一种新的模型提供了遗传学证据，在该模型中，表面外胚层必须接收到BMP信号，导致FGFs下调，进而诱导潜在的间充质细胞凋亡(Pajni-Underwood S.等人2007年开发134：2359)。因此，我们证明BMPs通过调节成纤维细胞生长因子信号间接控制细胞程序性死亡。我们通过研究BMP和FGF信号在肢体发育的各个方面的作用，利用在发育肢体特定区域表达Cre的小鼠品系，扩展了这些研究。例如，检验BMPs作为细胞死亡直接效应因子的假设的唯一方法是，仅在经历细胞死亡的谱系中灭活BMPs受体，而不影响附近细胞中的FGF表达。我们已经使用新的Cre品系实现了这一点，这种品系允许Cre介导的基因在这些谱系中失活。这些问题是：BMP是正常的程序性细胞死亡的直接影响因素吗？如果不是，程序性细胞死亡是如何控制的？如果是这样，BMP是如何实现这一目标的？在另一项研究中，我们发现了FGFs和BMP之间的一个重要信号节点，它对肢体骨骼的正常发育至关重要。我们之前的工作，如上所述，证明了特定的FGFs，从肢芽的远端结构中分泌出来，调节肢体的正常生长和模式。在目前的工作中，我们正在产生遗传学证据，表明BMP向骨骼元素的前体群体发出信号，通过控制成纤维细胞生长因子拮抗剂的表达来调节这种成纤维细胞生长因子信号。这两个信号通路的连接不仅是对肢体如何形成模式的独特洞察，而且可能为这两个通路在其他发育背景下或癌症期间如何相互作用提供一个模型。

One fundamental problem in development is to determine how cells decide to remain multipotent or to differentiate. This problem lies at the heart of how pattern is generated in the embryo and, conversely, how pattern is disrupted when these processes are damaged as in tumor formation. The developing limb is an ideal system to address these problems because a wealth of data exists concerning limb development due to studies in genetics and experimental embryology. Previously, we have shown an essential role for Fibroblast Growth Factor (FGF) signaling during limb development by examining mice lacking genes that encode FGF ligands (Lewandoski et al. 2000 Nature Genetics 28:167, Sun et al 2000 Nature Genetics 25: 6 ). However, the complexity caused by Fgf gene redundancy has led us to consider approaching the problem by examining FGF receptor mutants leading to the insight that FGF signaling controls limb size (Verheyden et al 2005Development 132:4235). Signaling through the pathway governed by Bone Morphogenetic Proteins (BMPs) is thought to play a role in all aspects of limb outgrowth: early patterning in all three axes, programmed cell death and bone formation. The task before us is to understand how BMP and FGF signaling pathways interact during limb development. One process that BMP are thought to play a direct role in is as effectors of normal programmed cell death that occurs in mesenchymal interdigit cells, thus removing them and sculpting the final digit pattern in animals that are born without webbed limbs. However, we have challenged this paradigm in our recent publication (Pajni-Underwood S. et al 2007 Development 134: 2359). By simultaneously inactivating the Bmp receptor gene, Bmpr1a as well as Fgf8 and Fgf4 specifically in the limb bud ectoderm, we have produced genetic evidence for a novel model in which the surface ectoderm must receive a BMP signal, resulting in down regulation of Fgfs which in turn induces apoptosis of the underlying mesenchyme (Pajni-Underwood S. et al 2007 Development 134: 2359). Thus we demonstrated that BMPs control programmed cell death indirectly, by regulating FGF signaling. We have extended these studies by studying the role of BMP and FGF signaling in various aspects of limb development using mouse lines that express Cre in specific region of the developing limb. For example the only way to test the hypothesis that BMPs act as direct effectors of cell death is to inactivate BMPs receptors only in the lineage that undergoes cells death, without affecting FGF expression in nearby cells. We have achieved this using new Cre lines that allow Cre-mediated gene inactivation in these lineages. With these lines are asking: are BMPs are direct effectors of normal programmed cell death? If not, how is programmed cell death controlled? If so, how do BMPs achieve this endpont? In another study, we have uncovered an important node of signaling between FGFs and BMP that is essential for normal development of the limb skeleton. Our previous work, cited above, demonstrates that specific FGFs, secreted from a distal structure in the limb bud, regulate the normal outgrowth and patterning of the limb. In current work, we are generating genetic evidence that BMP signaling to the progenitor population of the skeletal elements regulates this FGF signal by controlling the expression of an FGF antagonist. This linking of the two signaling pathways is not only a unique insight into how the limb is patterned but may provide a model for how the two pathways interact in other developmental contexts or during cancer.