Ethanol-induced disruption of kinase signaling pathways in brain development

乙醇诱导大脑发育中激酶信号通路的破坏

• 批准号: 10366867
• 负责人: ERIC Christopher OLSON
• 金额: $ 36.68万
• 依托单位: UPSTATE MEDICAL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-19 至 2027-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10366867
• 关键词: Actins; Acute; Adaptor Signaling Protein; Alcohols; Axon; Biochemical; Biological; Brain; Cells; Centers for Disease Control and Prevention (U.S.); Child; Critical Pathways; Defect; Dendrites; Development; Disease; Dissection; Dose; Elements; Embryo; Enzymes; Ethanol; Ethanol dependence; Etiology; Event; Fetal Alcohol Exposure; Fetal Alcohol Syndrome; Future; Genetic; Genetic Predisposition to Disease; Genetic study; Goals; Golgi Apparatus; Growth; In Vitro; Intellectual functioning disability; Knock-out; Ligand Binding; Ligands; Live Birth; Location; Membrane; Methodology; Molecular; Molecular Target; Morphology; Multipolar Neuron; Mus; Nervous system structure; Neuronal Differentiation; Neurons; Phase; Phosphorylation; Phosphotransferases; Positioning Attribute; Protein Dephosphorylation; Protein Tyrosine Kinase; Proteins; Receptor Protein-Tyrosine Kinases; Reelin Signaling Pathway; Research Personnel; Role; Signal Pathway; Signal Transduction; Site; Structure; Synaptic plasticity; Syndrome; Teratogenic effects; Testing; Tyrosine Phosphorylation; United States; alcohol consumption during pregnancy; alcohol effect; alcohol exposure; alcohol research; alcohol response; axon growth; axonal pathfinding; cofilin; fetal; glycosylation; in vivo; maternal alcohol use; mind control; novel; postnatal; prevent; receptor; receptor binding; response; src-Family Kinases; synaptic function; white matter

ABSTRACT Fetal Alcohol Syndrome (FAS) is one of the leading causes of intellectual disability in the United States. The CDC estimates that 0.2-1.5 per 1000 live births are children with FASD, a syndrome characterized by disrupted fetal brain development and postnatal intellectual disability (ID). Disrupted connectivity including altered dendritic structure, axonal pathfinding and white matter tracts are common findings in FAS and are thought to be major contributors to ID. However, the cellular and biological targets of alcohol are diverse and it is not clear whether there are common underlying molecular mechanisms producing these disruptions. Identification of common molecular mechanism(s) would enable a deeper understanding of this disorder, inform studies of genetic susceptibilities and provide molecular targets for neuroprotective strategies. This proposal pursues our finding that acute ethanol (EtOH) exposure disrupts Src kinase activity in embryonic cortical neurons. Src is a critical non-receptor tyrosine kinase that sits at central positions in multiple signaling pathways including the Reelin-Dab1 signaling pathway which controls brain layer formation and dendritogenesis. We found that acute EtOH exposure activates Src and induces phosphorylation of many proteins including Dab1, an essential adaptor protein in the Reelin-signaling pathway. Remarkably, this dramatic increase in phosphorylation is followed by a sustained dephosphorylation response in which the phosphorylation of Reelin effectors including Dab1, Src itself and the actin severing protein n-cofilin return to baseline levels, or below. During the extended dephosphorylation phase, the Reelin-signaling pathway can no longer be activated by in vitro application of its ligand, Reelin. In AIM 1 of this proposal, we will determine whether Reelin-Dab1 silencing occurs in vivo after maternal dosing with EtOH. We will then determine whether genetic deficiency in Src prevents the phosphorylation and dephosphorylation responses. Genetically establishing the critical kinase that initiates the EtOH response in vivo will be essential for future neuroprotective efforts. We and others have shown that Reelin-Dab1 signaling controls Golgi-deployment in the forming dendrite. In AIM 2 we will examine whether Src activation and inactivation disrupts Golgi location and function. Disrupted Golgi function would be expected to impact membrane addition, glycosylation, secretion and appropriate expression of many proteins, with potential long term negative consequences on neuritogenesis and neuronal function. In AIM 3 we will determine whether the EphA3 signaling pathway is similarly disrupted by Src dysregulation. EphA3 is a receptor tyrosine kinase that is required for axonal and white matter tract development. We identified the activation site of EphA3 as a target of EtOH-induced Src dysregulation raising the possibility that EphA3 activation and then silencing may contribute to FASD-related white matter disruptions. Collectively, these studies will determine the contribution of EtOH-dependent Src dysregulation to altered developmental signaling in pathways critical for brain development.

摘要 胎儿酒精综合征（FAS）是美国智力残疾的主要原因之一。的 CDC估计，每1000名活产婴儿中有0.2 - 1.5名患有FASD，这是一种以破坏性发育为特征的综合征。 胎儿脑发育和出生后智力残疾（ID）。连接中断，包括改变 树突状结构、轴突寻路和白色物质束是FAS的常见发现， 然而，酒精的细胞和生物靶点是多种多样的，目前还不清楚。 是否有共同的潜在分子机制产生这些破坏。鉴定 共同的分子机制将使人们能够更深入地了解这种疾病， 遗传易感性并为神经保护策略提供分子靶点。这项建议追求我们的 发现急性乙醇（EtOH）暴露会破坏胚胎皮层神经元中Src激酶的活性。src是 一种重要的非受体酪氨酸激酶，位于多种信号通路的中心位置，包括 Reelin-Dab1信号通路控制脑层形成和树突状细胞生成。我们发现急性 EtOH暴露激活Src并诱导许多蛋白质的磷酸化，包括Dab 1，一种必需的蛋白质。 接头蛋白在Reelin信号通路。值得注意的是，这种磷酸化的急剧增加， 随后是持续的去磷酸化反应，其中包括 Dab1、Src本身和肌动蛋白切割蛋白n-cofilin恢复到基线水平或更低。在延长 在去磷酸化阶段，Reelin信号通路不能再通过体外应用其 配体，Reelin。在本建议的AIM 1中，我们将确定在体内是否发生Reelin-Dab1沉默， 母体给药EtOH。然后我们将确定Src基因缺陷是否会阻止 磷酸化和去磷酸化反应。从遗传学上建立起启动 体内EtOH反应将是未来神经保护工作的关键。我们和其他人已经证明， Reelin-Dab1信号控制高尔基体在树突形成中的部署。在AIM 2中，我们将检查Src是否 激活和失活破坏了高尔基体的位置和功能。高尔基体功能的破坏 影响膜添加、糖基化、分泌和许多蛋白质的适当表达， 对轴突发生和神经元功能的潜在长期负面后果。在AIM 3中， 确定EphA3信号通路是否类似地被Src失调破坏。EphA3是一种 轴突和白色物质束发育所需的受体酪氨酸激酶。我们确定了 EphA3的激活位点作为EtOH诱导的Src失调的靶点，这提高了EphA3 激活然后沉默可能导致FASD相关的白色物质破坏。总的来说，这些 研究将确定EtOH依赖性Src失调对发育信号改变的贡献 大脑发育的关键途径。