Characterizing the mechanistic basis for sex-dimorphic responses to retinoic acid signaling in the developing brain

表征发育中大脑对视黄酸信号的性别二态性反应的机制基础

• 批准号: 10607935
• 负责人: Kelsey Hennick
• 金额: $ 4.36万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-06-30 至 2025-06-29
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10607935
• 关键词: Address; Anatomy; Anterior; Basic Science; Biological Assay; Biomedical Research; Brain; CRISPR interference; Central Nervous System; Data; Data Set; Development; Disease; Disparate; Enhancers; Enzymes; Epigenetic Process; Etiology; Exhibits; Exposure to; FOXP1 gene; Female; Fragile X Syndrome; Gene Dosage; Gene Expression; Gene Expression Regulation; Genes; Genetic; Genetic Transcription; Genomics; Glutamates; Gonadal Steroid Hormones; Histones; Human; Immunohistochemistry; In Vitro; Incidence; KDM5B gene; Lead; Link; Lysine; Mediating; Methylation; Modeling; Molecular; Mutation; Neurodevelopmental Disorder; Neurons; Organoids; Pattern; Penetrance; Phenotype; Prefrontal Cortex; RORA gene; Regulator Genes; Retinoic Acid Receptor; Rett Syndrome; Sampling; Sex Bias; Sex Chromosomes; Sex Differences; Signal Pathway; Signal Transduction; Site; Structure; Testing; Therapeutic Intervention; Tissues; Transcription Initiation; Transcriptional Regulation; Tretinoin; Up-Regulation; Vitamin A; Work; X Chromosome; X Inactivation; autism spectrum disorder; cell type; demethylation; differential expression; dosage; experimental study; genome-wide; gray matter; histone modification; human female; insight; knock-down; male; morphogens; nervous system development; nervous system disorder; neurodevelopment; programs; receptor function; response; retinoic acid receptor alpha; risk variant; sex; sexual dimorphism; single-cell RNA sequencing; stem cell differentiation; stem cells; targeted treatment; therapeutic candidate; transcription factor

Project Summary/Abstract Sex hormones and sex chromosome genes instruct development of sex-specific features during neurodevelopment, including differences in somatic gene expression and gray matter volume. Together, these developmental programs lead to fundamental differences at the molecular, cellular, and tissue level between males and females. While some neurological disorders have clear links to X chromosome genes, such as Rett Syndrome and Fragile X Syndrome, other disorders including Autism show sex-biased penetrance with no clear genetic mechanism. This proposal aims to address the mechanistic basis of sex-dimorphic transcriptional responses to retinoic acid signaling in the developing brain. Retinoic acid signaling is indispensable for modulating gene regulatory programs that orchestrate proper nervous system development, and recent work has shown that the Autism risk gene FOXP1 is upregulated in response to retinoic acid signaling. Preliminary studies have shown specific upregulation of FOXP1 in response to retinoic acid signaling in female cortical organoids, and not in males. This project will similarly leverage stem cell-derived cortical organoids to model neurodevelopment in vitro to better understand the molecular basis of sex-dimorphic phenotypes upon exposure to retinoic acid. Aim 1 will characterize sex-dimorphic genome wide expression changes in response to retinoic acid. Male and female organoids will be treated with vitamin A, the precursor to retinoic acid, and used for scRNA-seq. This dataset will uncover genes upregulated specifically in female organoids in response to retinoic acid. Additionally, I will identify cell types in both male and female organoids that exhibit the greatest gene expression changes in response to retinoic acid, lending insight into cell type-specific sensitives to retinoic acid during neurodevelopment. In Aim 2, the relationship between dosage of X-linked lysine demethylase KDM5C and retinoic acid-induced sex-specific gene regulatory programs will be characterized. Knockdown of KDM5C in female stem cell-derived organoids and subsequent genome-wide changes in H3K4 methylation, which is demethylated by KDM5C, will be determined by CUT&Tag. A putative enhancer at the FOXP1 locus coincides with H3K4me3 marks, and KDM5C-dependent methylation at this genomic site will be of particular importance. The impact of KDM5C knockdown on retinoic acid-induced FOXP1 expression will then be interrogated by immunohistochemistry. Together, these experiments will further our understanding of the intersection of epigenetics, gene expression, and cell signaling pathways during neurodevelopment, providing an an important mechanistic basis for sex-dimorphic developmental programs. This will further our understanding of the etiology of neurodevelopmental disorders with sex-biased penetrance, while uncovering potential candidates for therapeutic interventions.

项目总结/摘要 性激素和性染色体基因指导性特异性特征的发育， 神经发育，包括体细胞基因表达和灰质体积的差异。所有这些 发育程序导致了分子、细胞和组织水平上的根本差异， 雄性和雌性。虽然一些神经系统疾病与X染色体基因有明确的联系， 综合征和脆性X综合征，包括自闭症在内的其他疾病显示出性别偏见， 明确的遗传机制。该建议旨在解决性别二态转录的机制基础， 在发育中的大脑中对视黄酸信号的反应。视黄酸信号是必不可少的， 调节基因调控程序，协调适当的神经系统发育，以及最近的工作， 研究表明，自闭症风险基因FOXP 1在响应视黄酸信号时上调。初步 研究表明，在雌性皮质中，FOXP 1响应于视黄酸信号传导而特异性上调， 类器官，而不是在男性。该项目将同样利用干细胞衍生的皮质类器官来建模 体外神经发育，以更好地了解性别双态表型的分子基础， 接触视黄酸。目的1将描述性别二态性基因组范围内的表达变化， 视黄酸。男性和女性类器官将用维生素A（维甲酸的前体）治疗， 用于scRNA-seq.该数据集将揭示女性类器官中特异性上调的基因， 视黄酸。此外，我将确定男性和女性类器官中表现出最大的细胞类型。 基因表达的变化，响应视黄酸，贷款洞察细胞类型特异性敏感， 视黄酸在神经发育中的作用在目标2中，X-连接的赖氨酸的剂量与 脱甲基酶KDM 5C和视黄酸诱导的性别特异性基因调控程序的特点。 女性干细胞衍生的类器官中KDM 5C的敲低以及随后H3 K4的全基因组变化 由KDM 5C去甲基化的甲基化将由CUT&Tag测定。一个假定的增强子， FOXP 1基因座与H3 K4 me 3标记一致，并且该基因组位点的KDM 5C依赖性甲基化将被 特别重要。KDM 5C敲低对视黄酸诱导的FOXP 1表达的影响将 然后通过免疫组织化学进行询问。总之，这些实验将进一步加深我们对 神经发育过程中表观遗传学、基因表达和细胞信号传导途径的交叉， 为性别二态发育程序提供了重要的机制基础。这将进一步促进我们的 了解神经发育障碍的病因与性别偏见的恍惚，同时揭示 治疗干预的潜在候选者。