LIM-HD/bHLH Combinatorial Code in Motoneurons

我们的长期目标是了解发展中的组合转录调控网络， 中枢神经系统（CMS），其在指定大量不同的神经元和 神经胶质细胞类型。LIM同源结构域（LIM-HD）和碱性螺旋-环-螺旋（bHLH）蛋白质是相对较好的 特征转录因子调节细胞类型规范在胚胎CMS。但无论是 这些因子调节基因表达的分子机制及其靶基因的特性 仍然知之甚少。我们希望探讨这些重要问题，具体侧重于它们在以下方面的作用： 产生脊髓运动神经元（MN）。我们的结果表明，LIM-HD因子Lhx 3和Isl 1指定 胚胎脊髓的两个不同的神经元以组合的方式。Lhx 3和LIM辅因子NLI（对于 核LIM-相互作用子）形成V2-中间神经元（IN）-特异性四聚体。相比之下，Lhx 3和NLI形成了一个 MN-特异性六聚体与Isl 1共表达。在MN特化期间，bHLH蛋白Ngn 2/NeuroM 在功能上与六聚体同步，尽管这种新的分子机制 串扰没有被完全理解。我们最近的筛选工作导致发现了具有 NLI：lsl 1：Lhx 3六聚体结合位点，其中许多与候选MN基因相连。这些结果导致我们 该建议的中心假设：六聚体直接调节多个MN基因与bHLH 影响MN规格的因素。我们将剖析这一假设，以揭示分子基础， MN特化响应于转录因子的组合表达。具体而言，研究 提出探索六聚体/bHLHs与其候选靶基因的不同功能方面。 总的来说，拟议的研究应提供关键的见解，为正确形成的基本原则， 神经系统和治疗各种神经元损伤和疾病的实用信息。值得注意的是， LIM-HD/bHLH因子还参与指定发育中CNS的其他区域中的细胞命运（例如， 视网膜和前脑）以及非中枢神经系统器官，如心脏，胰腺和垂体，这突出了 建议研究的重要性。

Our long-term goal is to understand the combinatorial transcriptional regulatory networks in the developing central nervous system (CMS), which play essential roles to specify a large number of distinct neuronal and glial cell types. LIM homeodomain (LIM-HD) and basic helix-loop-helix (bHLH) proteins are relatively well characterized transcription factors regulating cell type specification in the embryonic CMS. However, both the molecular mechanism by which these factors regulate gene expression and the identity of their target genes remain poorly understood. We wish to explore these important issues by specifically focusing on their role in generating spinal motoneurons (MNs). Our results demonstrate that LIM-HD factors Lhx3 and Isl1specify two distinct neurons of the embryonic spinal cord in a combinatorial manner. Lhx3 and the LIM cofactor NLI(for nuclear LIM-interactor) form a V2-interneuron (IN)-specifying tetramer. In contrast, Lhx3 and NLI form a MN-specifying hexamerwhen coexpressed with Isl1. During MN specification, bHLH proteins Ngn2/NeuroM are functionally synchronized with the hexamer, although the molecular mechanism underlying this novel crosstalk is not fully understood. Our recent screening effort led to the finding of genomic fragments with the NLI:lsl1 :Lhx3 hexamer binding sites, many of which are linked to candidate MN genes. These results led to our central hypothesis of this proposal: the hexamer directly regulates multiple MNgenes in conjunction with bHLH factors to effect MN specification. We will dissect this hypothesis to uncover the molecular basis that directs MN specification in response to a combinatorial expression of transcription factors. Specifically, studies are proposed to explore the diverse functional aspects of the hexamer/bHLHs with their candidate target genes. Overall, the proposed studies should provide critical insights into the basic principles for the proper formation of the nervous system and practical information to treat a variety of neuronal injuries and diseases. Of note, LIM-HD/bHLH factors are also involved with specifying cell fates in other regions of the developing CNS (e.g., retina and forebrain) as well as non-CNS organs such as heart, pancreas and pituitary, which highlights the general importance of the proposed studies.