Isoform expression and post-transcriptional regulation of centrosomal plp mRNA

中心体 plp mRNA 的亚型表达和转录后调控

• 批准号: 10646408
• 负责人: Junnan Fang
• 金额: $ 12.5万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-01 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10646408
• 关键词: 3' Untranslated Regions; Award; Bioinformatics; Biological; Biological Assay; Biological Process; Biology; Brain; Cardiovascular Diseases; Cell physiology; Centrioles; Centrosome; Cephalic; Clustered Regularly Interspaced Short Palindromic Repeats; Congenital Disorders; Coupled; Data; Defect; Development; Disease; Down Syndrome; Down-Regulation; Drosophila genus; Embryo; Etiology; Gene Expression; Gene Expression Regulation; Generations; Genes; Health; Human; Individual; Knowledge; Laboratories; Length; Male Sterility; Male Sterilizations; Malignant Neoplasms; Mass Spectrum Analysis; Mentors; Messenger RNA; Microcephaly; Microtubule-Organizing Center; Microtubules; Modeling; Molecular Weight; Monitor; Mutate; Neurodegenerative Disorders; Neurodevelopmental Disorder; Neuronal Dysfunction; Orthologous Gene; Pattern; Phase; Phenotype; Poly A; Poly(A) Tail; Polyadenylation; Post-Transcriptional Regulation; Protein Isoforms; Proteins; RNA; RNA Splicing; RNA-Binding Proteins; Regulation; Reporter; Research Personnel; Role; Running; Structure; Technical Expertise; Testing; Testis; Tissues; Training; Training Programs; Transcript; Translations; Up-Regulation; Variant; Western Blotting; Work; cell motility; cell type; ciliopathy; developmental disease; differential expression; genome editing; human disease; imaging approach; improved; insight; neuroblast; neurogenesis; osteodysplastic primordial dwarfism; pericentrin; programs; promoter; protein function; recruit; scaffold; spatiotemporal; transcriptome sequencing; transcriptomics

PROJECT SUMMARY Centrosome deregulation is associated with developmental disorders, such as microcephaly, ciliopathy, and cardiovascular disease. Despite their fundamental importance to human health, relatively little is known about the regulation of genes encoding core centrosome components, such as Pericentrin (Pcnt)-like protein (PLP), a conserved centrosome scaffold also required for ciliary function. Completion of this proposal will advance our understanding of the post- transcriptional regulation of plp mRNA. In humans, deregulation of the homologous PCNT gene results in congenital diseases, such as microcephalic osteodysplastic primordial dwarfism type II (MOPD II) and Trisomy 21. Similarly, in Drosophila, plp deregulation leads to diverse defects, including embryonic lethality, neuron dysfunction, and male sterility. The mechanisms underlying the pleiotropic phenotypes associated with plp loss are incompletely understood. plp is predicted to encode 12 mRNA variants, but what mechanisms give rise to these distinct RNA species and how their expression is spatiotemporally regulated are completely unknown. In this K99/R00 proposal, I will test the hypothesis that the PLP protein isoform expression, coupled with the post- transcriptional regulation of plp mRNA, modulates its diverse functions within different tissues. Three aims are proposed to test this hypothesis. In Aim 1, I will identify mechanisms of embryonic plp mRNA localization and translation. In Aim 2, I will explore the contribution of alternative promoter and 3’UTR usage for the generation of different plp RNA variants. In Aim 3, I will determine the expression profile of PLP protein isoforms and examine the biological function of PLP isoforms, including PLPPM, in Drosophila neuroblasts versus early embryos. The completion of this work will reveal the mechanisms of spatially and temporally distinct expression patterns of functional PLP protein isoforms in different Drosophila tissues and will improve our understanding of plp mRNA regulation, aspects of which may be deregulated in human diseases, such as neurodegenerative disorders and cardiovascular disease. Moreover, this award will provide technical and professional training in RNA- sequencing and bioinformatics, CRISPR genome editing, and advanced imaging approaches under the guidance of my expert mentors. I will follow a structured training program to enhance my professional abilities to establish and run my own successful independent laboratory.

项目摘要 中心体放松管制与发育障碍有关，例如小头畸形，纤毛病和 心血管疾病。尽管它们对人类健康的重要性至关重要，但鲜为人知 关于编码核心中心体成分的基因的调节，例如包膜蛋白（PCNT）样蛋白 （PLP），睫状功能也需要保守的中心体支架。该提议的完成将 促进我们对PLP mRNA的转录后调节的理解。在人类中，放松管制 同源PCNT基因会导致先天性疾病，例如微头性骨质增生性疾病 原始矮人II型（MOPD II）和三体疾病21。同样，在果蝇中，PLP放松管制导致 各种缺陷，包括胚胎致死性，神经元功能障碍和雄性不育。机制 与PLP损失相关的多效性表型的基础是未完全理解的。预测PLP 编码12个mRNA变体，但是哪些机制产生了这些不同的RNA种类以及它们如何 表达在空间上是完全未知的。在此K99/R00建议中，我将测试 PLP蛋白同工型表达，与PLP的转录后调控相结合的假设 mRNA，调节其在不同组织中的潜水功能。提出了三个目标来测试这一点 假设。在AIM 1中，我将确定胚胎PLP mRNA定位和翻译的机制。在AIM 2中， 我将探讨替代启动子和3'UTR使用的贡献，以生成不同的PLP RNA 变体。在AIM 3中，我将确定PLP蛋白同工型的表达曲线并检查生物学 果蝇神经细胞与早期胚胎中PLP同工型（包括PLPPM）的功能。完成 这项工作将揭示空间和暂时不同的功能表达模式的机制 不同果蝇组织中的PLP蛋白同工型，将提高我们对PLP mRNA的理解 调节，其各个方面可能在人类疾病中放松管制，例如神经退行性疾病 和心血管疾病。此外，该奖项将提供RNA的技术和专业培训 - 测序和生物信息学，CRISPR基因组编辑和高级成像方法 我的专家导师的指导。我将遵循结构化的培训计划，以增强我的专业能力 建立并运营自己成功的独立实验室。