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。该过程开始于 神经祖细胞沿背腹轴和前后轴沿着形成图案，以产生不同的亚型， 神经元许多关键转录因子（TF）控制神经元亚型特化的过程。工作 小鼠的研究表明，同源域（HD）TF Gsx 2在细胞的模式化和分化中起着重要作用。 由胚胎小鼠外侧神经节隆起（LGE）中的祖细胞产生的神经元细胞类型 端脑这些祖细胞产生的细胞类型包括基底神经节的纹状体投射神经元 和嗅球中的中间神经元，这两者在小鼠Gsx 2突变体中都严重减少。因此，人类 患者研究在具有严重神经系统症状的儿童中确定了2种病理性GSX 2变异等位基因，包括 肌张力障碍和智力残疾。与这些症状一致，MRI成像显示严重的基底神经节 发育不全一个GSX 2变体导致无效等位基因，然而，另一个是错义变体（Q251 R），其改变了一个关键的 DNA中的氨基酸结合HD。我们建立了这种人类变异的小鼠模型，我们的初步研究表明， Q>R变异导致了一种强的胚胎LGE和基底神经节表型，其在形态上类似于 Gsx 2无效等位基因的胚胎。此外，我们的初步数据表明，这种人类HD变体改变了Gsx 2 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Gsx 2 通过两种机制结合和调节靶基因;作为单体，Gsx 2抑制基因表达，而在子集上， Gsx 2与二聚体位点的协同结合似乎有利于基因表达。有趣的是，Dlx HD 在LGE祖细胞成熟过程中位于Gsx 2下游的TF也结合单体位点，但不是抑制Gsx 2的表达。 它们激活基因表达。在本申请中，我们提出确定Gsx 2和Dlx TF如何调节LGE 基底神经节发育期间的基因表达。为了实现这一目标，我们将在3中测试以下假设 独立的具体目的：1）检验Gsx 2通过介导不同的细胞因子来控制基底神经节发育的假设。 以DNA结合位点依赖的方式调控基因的结果。2)为了检验Gsx 2和Dlx TF 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)为了检验这一假设 GSX 2 Q251 R人类变异体导致DNA结合特异性改变，从而导致 LGE基因表达和最终基底神经节发育不全。我们的方法将联合收割机结合使用小鼠遗传学和 用分子、生物化学和基因组方法研究人前脑神经干细胞的转录 控制发育中的基底神经节的神经元特化。CCHMC研究团队的独特专业知识 允许我们采用这种广泛的方法，从而增加了我们深入了解Gsx因素的机会 控制基底神经节的发育，并揭示新的基因调控机制， 某些儿童神经系统疾病