Patterning of late-acting germinal zones in the vertebrate CNS

描述(由申请人提供)：神经科学和再生医学的一个基本挑战是了解特定的神经元亚型如何在正确的数量、确定的时间和精确的位置产生。例如，通过对早期脊髓神经管的研究，已经开始确定协调这类事件的计划；部分原因是物理和分子上的可及性，以及相对简单的解剖学--新生的神经元通常径向移动较短距离，维持原始的DV/AP关系。不太清楚的是，这类事件是如何在生发区受到调控的，生发区通过切向迁移模式将神经元队列分布到遥远和不同的位置，而且通常在发育后期这样做。然而，这种生发带对大脑中神经元的多样性有很大贡献，而且鉴于它们的独特性质和妊娠后期神经组织的独特环境，必须以新的方式做到这一点。这项提议试图通过对后脑(下部)菱形唇(LRL)的研究来填补这一知识空白的关键方面，LRL是一种迟起作用的基本生发区，现在可以通过分子遗传学手段获得。LRL产生了多种不同的、功能关键的脑干细胞类型，其中许多细胞容易发生发育和退行性疾病。我们提供的证据表明：(1)产生的细胞类型的这种多样性反映在精确定义的LRL的分子亚区域化中-LRL不是同质的，而是由预测后代神经元身份的DV和AP基因表达微域组成；(2)转录因子Pax6调节这种分子亚区域化的各个方面，确保产生正确比例的后代谱系；以及(3)原型的腹腔化形成因子Sonic hedgehog(Shh)在位于背部的后脑脉络丛上皮、已知的背侧组织中心和LRL谱系中表达。这种先前排除的构图分子，如Pax6和Shh的共同选择，可能代表了一种总体策略，通过这种策略，晚作用的生发区可能会继续产生独特的祖细胞状态和命运。我们的目标是双重的：通过将高分辨率的DV：AP坐标与后来产生的细胞命运联系起来，扩展我们的LRL命运图(目标1)；在这个图上放置调节这种分子组织的分子的作用，从而产生特定的细胞类型；例如，我们将探索Pax6和Shh(目标2和3)的新角色。

DESCRIPTION (provided by applicant): A fundamental challenge in neuroscience and regenerative medicine is to understand how specific neuron subtypes arise in correct number, at defined times, and target to precise locations. Programs enacted to coordinate such events have begun to be defined, for example, through studies of early-stage spinal neural tube; in part due to physical and molecular accessibility, and relatively simple anatomy--newly born neurons typically move short distances radially, maintaining originating DV/AP relationships. Less clear is how such events are regulated in germinal zones that distribute neuron cohorts to distant and varied locations through tangential migration modes, and which typically do so at later developmental stages. Yet such germinal zones contribute substantially to neuron diversity in the brain and must do so in novel ways given their unique properties and the distinct milieu of late-gestation neural tissue. This proposal seeks to fill key aspects of this knowledge gap through studies of the hindbrain (lower) rhombic lip (LRL), a late-acting and essential germinal zone now accessible by molecular genetic means. Diverse and functionally critical brainstem cell types arise from the LRL, many subject to developmental and degenerative disorders. We provide evidence that (1) this diversity in produced cell types is reflected in a precisely defined molecular sub-regionalization of the LRL - the LRL is not homogeneous, but rather comprised of DV and AP gene expression microdomains predictive of progeny neuron identity; (2) the transcription factor Pax6 modulates aspects of this molecular sub-regionalization, ensuring that correct proportions of descendant lineages are produced; and (3) the archetypal ventralizing morphogen, Sonic hedgehog (Shh), is expressed in the dorsally situated hindbrain choroid plexus epithelium, a known dorsal organizing center and itself a LRL lineage. Such co-opting of previously excluded patterning molecules, such as Pax6 and Shh, may represent a general strategy by which a late-acting germinal zone may continue generating unique progenitor cell states and fates. Our goal is to two-fold: Extend our LRL fate map by linking highly resolved DV:AP coordinates to later-generated cell fates (Aim 1); place on this map the action of molecules which regulate this molecular organization and thus production of specific cell types; for example, we will explore novel roles for Pax6 and Shh (Aims 2 & 3).