DRG Progenitor: Role of extrinsic and intrinsic cues

描述（由申请人提供）：周围神经系统（PNS）是生命所必需的。我们对氧气水平、二氧化碳、血压的变化做出自主反应的能力，以及对可能伤害生物体的有害刺激的检测，都是由PNS介导的关键功能。当这些系统的发育出现问题时，就会导致感觉和/或自主神经病变，包括隐性遗传病家族性自主神经异常（FD），在这种疾病中，感觉和自主神经系统都不能正常发育。大多数PNS来源于一群不切实际的干细胞，即神经嵴。这些细胞从神经管脱落，并在整个胚胎中沿着固定的轨迹迁移，最终在PNS内产生大多数衍生物。虽然在过去的10年里，许多介导感觉神经元发育的分子机制已经被确定，但我们对协调NCCs产生特定衍生物的行为的细胞机制的理解很少。然而，随着实时成像技术和荧光蛋白变异试剂的革命，再加上对禽类胚胎进行体内增益和损失函数扰动的容易性，现在可以在进行分子扰动的同时实时成像神经嵴细胞的迁移和分化。我们将结合这些强大的技术来研究不同的神经嵴细胞亚群是否会在背根神经节中产生亚型特异性的感觉神经元。为此，我们将使用光激活GFP变异体、基因特异性报告构建体和逆转录病毒来追踪从神经管空间离散区域迁移的神经嵴细胞，跨越三个时间上不同的迁移波，并追踪它们的谱系，因为它们产生祖细胞和感觉神经元的亚型。此外，我们将扩展我们从研究正常感觉神经元发育中所学到的知识，通过分析小鼠和鸡的感觉神经元发育，研究导致FD的潜在分子和细胞机制，其中负责FD疾病的基因被删除（小鼠）或敲低（鸡）。

DESCRIPTION (provided by applicant): The peripheral nervous system (PNS) is essential for life. Our ability to autonomously respond to alterations in oxygen levels, CO2, blood pressure, and detection of noxious stimuli that could harm the organism, are all critical functions mediated by the PNS. When the development of these systems go awry, sensory and/or autonomic neuropathies result including the recessive genetic disease, Familial Dysautonomia (FD), in which neither the sensory nor the autonomic nervous systems develop correctly. Most of the PNS derives from a quixotic population of stem-like cells, the neural crest. These cells delaminate from the neural tube, and migrate along stereotyped trajectories throughout the embryo to ultimately give rise to the majority of derivatives within the PNS. Although over the past 10 years, many of the molecular mechanisms that mediate sensory neuron development have been identified our understanding of the cellular mechanisms that orchestrate the behaviors of NCCs as they give rise to specific derivatives is sparse. However, with the revolution in live imaging technologies and fluorescent protein variant reagents, combined with the ease of conducting in vivo gain and loss function perturbations in the avian embryo, it is now possible to image in real time the migration and differentiation of neural crest cells while simultaneously conducting molecular perturbations. We will combine these powerful technologies in this proposal to investigate whether distinct subpopulations of neural crest cells give rise to subtype-specific classes of sensory neurons in the dorsal root ganglia. To this end, we will use photoactivatable GFP variants, gene-specific reporter constructs, and retroviruses to track neural crest cells that emigrate from spatially discrete regions of the neural tube, over the three temporally distinct waves of emigration, and trace their lineage as they give rise to subtypes of progenitor cells and sensory neurons. Furthermore we will extend what we learn from studying normal sensory neuron development to investigate the underlying molecular and cellular mechanisms that go awry to result in FD by analyzing sensory neuron development in both mice and chick in which the gene responsible for the FD disease is deleted (mouse) or knocked-down (chick). PUBLIC HEALTH RELEVANCE: Our work is focused on understanding how pain-sensing neurons are born and mature. These neurons are essential for life as protection against noxious stimuli that could harm the organism. Achievement of the aims of our proposal will identify cellular and molecular mechanisms that can be applied to the treatment of both developmental and degenerative peripheral neuropathies including Familial dysautonomia and diabetic peripheral neuropathy.