The role of transcription factor SOX6 in midbrain dopamine neuron lineage and molecular diversity

转录因子SOX6在中脑多巴胺神经元谱系和分子多样性中的作用

• 批准号: 10437632
• 负责人: Maria Milagros Pereira Luppi
• 金额: $ 3.36万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-01 至 2022-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10437632
• 关键词: Adult; Affect; Anatomy; Animals; Biological; Cells; ChIP-seq; Classification; Comprehension; Conflict (Psychology); DNA; DNA Sequence; Data; Derivation procedure; Development; Disease; Disease model; Dopamine; Embryo; Event; Floor; Functional disorder; Gene Expression Regulation; Genes; Genetic; Genetic Transcription; Goals; Heterogeneity; In Vitro; Knock-out; Label; Lateral; Light; Maps; Medial; Methods; Midbrain structure; Modeling; Molecular; Molecular Target; Neurodegenerative Disorders; Neurons; Parkinson Disease; Population; Population Heterogeneity; Process; Protocols documentation; Publications; Regenerative Medicine; Resolution; Role; SOX6 gene; Substantia nigra structure; Testing; Time; base; cell replacement therapy; chromatin immunoprecipitation; cohort; design; dopaminergic neuron; experimental study; insight; motor control; nervous system disorder; neuron development; pars compacta; progenitor; recombinase; single-cell RNA sequencing; stem cells; therapy development; tool; transcription factor; transcriptome sequencing; transplant model

Dopamine (DA) neurons located in the Substantia nigra pars compacta (SNc) are involved in motor control, and their degeneration is associated with disorders such as Parkinson’s disease (PD), the second most frequent neurodegenerative disease in the US and practically incurable. Recent studies have revealed that DA neurons are molecularly, anatomically, and functionally heterogeneous. In fact, certain DA neuron subtypes are selectively vulnerable in animal PD models. Hence, recognizing this diversity will have a significant impact on the study of PD pathophysiology and the development of treatments. This proposal aims to shed light on early molecular events, particularly involving transcription factor SOX6, that govern DA neuron fate and diversification. All midbrain DA neurons originate from progenitors in the embryonic mesencephalic floor plate. Sox6 is expressed in progenitors of the most medial domain of the floor plate. It is widely accepted that Sox6-expressing progenitors give rise to the entire SNc DA neuron population, however, existing publications challenge this model suggesting that Sox6- progenitors, located in the lateral domain of the floor plate, also contribute to the adult SNc. Yet, none of these studies performed lineage analysis of Sox6. A proper Sox6 progenitor fate map would reconcile these results, however, existing lineage tracing tools are inadequate to resolve this progenitor-progeny relationship since Sox6 is expressed in progenitors and in postmitotic neurons. We have developed a new lineage tracing tool, a progenitor-restricted intersectional fate-mapping strategy (PRISM), that circumvents the limitations of recombinase-based strategies by anchoring the labeling to progenitors. With PRISM, we will unambiguously reveal the contribution of Sox6+ progenitors to specific DA neuron subtypes, particularly those found selectively vulnerable in PD models. Additionally, we will combine PRISM with a single-cell resolution analysis for the first time, to further refine the molecular landscape of the Sox6 progeny. Our lab has highlighted Sox6 expression as the distinctive variable in adult DA neuron classification. Still, more evidence is needed to establish whether SOX6 regulates genes that define specific neuronal subtypes. Exploring the role of Sox6 in development will help understand if progenitor pools are restricted to give rise to certain subtypes of the SNc. Additionally, elucidating the molecular targets of SOX6 will deepen our understanding of the determination of DA neuron fate. To this end, we will pair a Sox6 knockout model with RNA sequencing to determine the necessity of this gene in the diversification of DA neurons. We will also apply ChIP- Seq to find the main target DNA sequences of this transcription factor. The proposed aims will provide insights into the role of Sox6 in DA neuron development and diversification, with the hope of contributing to the optimization of in vitro protocols for regenerative medicine and disease modeling.

位于黑质致密部 (SNc) 的多巴胺 (DA) 神经元参与运动 它们的退化与帕金森病（PD）等疾病有关，帕金森病是第二大疾病 在美国，神经退行性疾病很常见，而且几乎无法治愈。最近的研究表明，DA 神经元在分子、解剖学和功能上是异质的。事实上，某些 DA 神经元亚型是 在动物 PD 模型中选择性易受影响。因此，认识到这种多样性将对 PD 病理生理学研究和治疗方法的开发。该提案旨在尽早阐明 分子事件，特别是涉及转录因子 SOX6，控制 DA 神经元的命运和多样化。 所有中脑 DA 神经元均起源于胚胎中脑底板的祖细胞。 Sox6 是 在底板最内侧区域的祖细胞中表达。人们普遍认为表达 Sox6 祖细胞产生整个 SNc DA 神经元群体，然而，现有出版物对这一模型提出了挑战 表明位于底板外侧区域的 Sox6- 祖细胞也有助于成年 SNc。然而，这些研究均未对 Sox6 进行谱系分析。正确的 Sox6 祖先命运图将 然而，为了协调这些结果，现有的谱系追踪工具不足以解决这个祖先-后代问题 由于 Sox6 在祖细胞和有丝分裂后神经元中表达，因此存在这种关系。我们开发了一种新的 谱系追踪工具，一种祖先限制的交叉命运映射策略（PRISM），可以规避 通过将标记锚定在祖细胞上，基于重组酶的策略的局限性。借助 PRISM，我们将 明确揭示 Sox6+ 祖细胞对特定 DA 神经元亚型的贡献，特别是那些 在 PD 模型中发现选择性脆弱。此外，我们将 PRISM 与单细胞分辨率相结合 首次分析，以进一步完善 Sox6 后代的分子景观。 我们的实验室强调 Sox6 表达是成人 DA 神经元分类中的独特变量。仍然， 需要更多证据来确定 SOX6 是否调节定义特定神经元亚型的基因。 探索 Sox6 在发育中的作用将有助于了解祖细胞库是否受到限制以产生 SNc 的某些亚型。此外，阐明 SOX6 的分子靶点将加深我们对 SOX6 的了解。 了解 DA 神经元命运的决定。为此，我们将 Sox6 敲除模型与 RNA 配对 测序以确定该基因在 DA 神经元多样化中的必要性。我们还将应用 ChIP- Seq 来查找该转录因子的主要靶标 DNA 序列。 拟议的目标将深入了解 Sox6 在 DA 神经元发育和 多样化，希望为再生医学和体外实验方案的优化做出贡献 疾病建模。