Roles of Gsx factors in basal ganglia development

Gsx 因子在基底神经节发育中的作用

• 批准号: 10339513
• 负责人: KENNETH J CAMPBELL
• 金额: $ 62.37万
• 依托单位: CINCINNATI CHILDRENS HOSP MED CTR
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-01 至 2026-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10339513
• 关键词: Alleles; Amino Acids; Anatomic Models; Anatomy; Anterior; Attention deficit hyperactivity disorder; Basal Ganglia; Behavioral; Binding; Binding Sites; Biochemical; Biochemistry; Bioinformatics; Biological Assay; Brain; Cell Culture Techniques; Cell Nucleus; Child; Childhood Neurological Disorder; Cognition; Corpus striatum structure; Coupled; DNA; DNA Binding; DNA-Binding Proteins; Data; Data Set; Development; Disease; Dorsal; Dystonia; Electrophoretic Mobility Shift Assay; Elements; Embryo; Enhancers; Exhibits; Functional disorder; Ganglia; Gene Activation; Gene Expression; Gene Expression Profiling; Gene Expression Regulation; Genes; Genetic; Genetic Enhancer Element; Genetic Transcription; Genomic approach; Genomics; Gilles de la Tourette syndrome; Goals; Grant; Histologic; Human; Image; In Vitro; Intellectual functioning disability; Interneurons; Lateral; Magnetic Resonance Imaging; Maps; Mediating; Modeling; Molecular; Morphology; Movement; Mus; Neurologic Symptoms; Neuronal Differentiation; Neurons; Nonsense Codon; Obsessive-Compulsive Disorder; Outcome; Pathologic; Patients; Pattern; Phenotype; Play; Process; Prosencephalon; Proteins; Publishing; Regulation; Regulator Genes; Regulatory Element; Reporter; Repression; Research; Resolution; Role; Site; Specificity; Symptoms; System; Telencephalon; Testing; Transcriptional Regulation; Variant; Work; behavioral phenotyping; cell type; dimer; functional genomics; genetic variant; homeodomain; human embryonic stem cell; in vivo; insight; monomer; mouse genetics; mouse model; mutant; nerve stem cell; neural circuit; neural patterning; neurogenesis; novel; olfactory bulb; postnatal; progenitor; transcription factor; transcriptome sequencing

Normal brain function relies on the correct assembly of neural circuits during development. This process starts with the patterning of neural progenitors along the dorsal-ventral and anterior-posterior axes to give rise to distinct subtypes of neurons. A number of key transcription factors (TFs) control the process of neuronal subtype specification. Work in the mouse has shown that the homeodomain (HD) TF Gsx2 plays essential roles in the patterning and differentiation of neuronal cell types that arise from progenitors in the lateral ganglionic eminence (LGE) of the embryonic mouse telencephalon. These progenitors give rise to cell types that include the striatal projection neurons of the basal ganglia and interneurons in the olfactory bulb, both of which are severely reduced in mouse Gsx2 mutants. Accordingly, human patient studies identified 2 pathological GSX2 variant alleles in children with serious neurological symptoms, including dystonia and intellectual disabilities. Consistent with these symptoms, MRI imaging revealed severe basal ganglia agenesis. One GSX2 variant results in a null allele, however, the other is a missense variant (Q251R) that alters a key amino acid in the DNA binding HD. We generated a mouse model of this human variant and our initial studies suggest that the Q>R variant leads to a strong embryonic LGE and basal ganglia phenotype that is morphologically similar to embryos with Gsx2 null alleles. Furthermore, our preliminary data indicate that this human HD variant alters Gsx2 DNA binding specificity, and thereby may account for the observed phenotypes. Moreover, we recently determined that Gsx2 binds and regulates target genes via two mechanisms; as a monomer Gsx2 represses gene expression whereas on a subset of DNA sites cooperative Gsx2 binding to dimer sites appears to facilitate gene expression. Intriguingly, the Dlx HD TFs, which lie downstream of Gsx2 during LGE progenitor maturation, also bind monomer sites but instead of repressing they activate gene expression. In this application, we propose to determine how Gsx2 and the Dlx TFs regulate LGE gene expression during basal ganglia development. To achieve this goal, we will test the following hypotheses in 3 independent specific aims: 1) To test the hypothesis that Gsx2 controls basal ganglia development by mediating distinct gene regulatory outcomes in a DNA binding site dependent manner. 2) To test the hypotheses that Gsx2 and Dlx TFs regulate a common set of LGE genes though direct competition for shared enhancer elements. 3) To test the hypothesis that the GSX2Q251R human variant causes altered DNA binding specificity, and thereby results in the mis-regulation of LGE gene expression and ultimately basal ganglia agenesis. Our approach will combine the use of mouse genetics and human forebrain neural stem cell cultures with molecular, biochemical, and genomic approaches to study transcriptional control of neuronal specification in the developing basal ganglia. The unique expertise of our research team at CCHMC allows us to take this broad approach, and thus increases our chances to gain a deeper understanding of how Gsx factors control basal ganglia development as well as to uncover new gene regulatory mechanisms that underlie dysfunction in certain childhood neurological disorders.

正常的大脑功能依赖于发育过程中神经回路的正确组装。此过程从 神经前体细胞沿背-腹轴和前-后轴的模式产生不同的亚型 神经元。许多关键的转录因子(TF)控制着神经元亚型的指定过程。在工作中工作 小鼠已经证明同源结构域(HD)Tf Gsx2在细胞的图案化和分化过程中起重要作用 胚胎小鼠外侧神经节隆起(LGE)祖细胞分化的神经细胞类型 端脑。这些祖细胞产生的细胞类型包括基底节的纹状投射神经元。 和嗅球中的中间神经元，这两个在小鼠Gsx2突变体中都严重减少。因此，人类 患者研究在有严重神经症状的儿童中发现了2个病理GSX2变异等位基因，包括 肌张力障碍和智力障碍。与这些症状相一致的是，核磁共振成像显示基底节严重 发育不全。然而，一个GSX2变体导致空等位基因，而另一个是改变密钥的错义变体(Q251R DNA结合HD中的氨基酸。我们制作了这种人类变种的小鼠模型，初步研究表明 Q&gt；R变异导致强烈的胚胎LGE和基底节表型，其形态类似于 具有Gsx2零等位基因的胚胎。此外，我们的初步数据表明，这种人类HD变体改变了Gsx2 DNA 结合特异性，从而可以解释观察到的表型。此外，我们最近确定Gsx2 Gsx2通过两种机制结合和调节靶基因；作为单体，Gsx2抑制基因表达，而在一个子集上 在DNA位点中，协同Gsx2与二聚体结合似乎促进了基因的表达。耐人寻味的是，DLX HD 在LGE前体成熟过程中，位于Gsx2下游的转录因子也与单体结合，但不是抑制 它们能激活基因表达。在本申请中，我们建议确定Gsx2和DLX TF如何调节LGE 基底节发育过程中的基因表达。为了实现这一目标，我们将在3中测试以下假设 独立的特定目标：1)检验Gsx2通过介导不同的 DNA结合位点依赖的基因调控结果。2)检验Gsx2和Dlx TF的假设 通过直接竞争共享的增强子元件来调节一组共同的LGE基因。3)检验假设 GSX2Q251R人类变异导致DNA结合特异性改变，从而导致错误调节 LGE基因的表达最终导致基底节发育不全。我们的方法将结合使用小鼠遗传学和 用分子、生化和基因组方法培养人前脑神经干细胞以研究转录 对发育中的基底节神经元规格的控制。我们CCHMC研究团队的独特专业知识 允许我们采取这种广泛的方法，从而增加了我们更深入地了解GSX因素的机会 控制基底神经节的发育，以及发现新的基因调控机制，这些机制是导致脑功能障碍的基础 某些儿童神经性障碍。