Isoform expression and post-transcriptional regulation of centrosomal plp mRNA

项目总结 中心体松弛与发育障碍有关，如小头症、纤毛病变和 心血管疾病。尽管它们对人类健康至关重要，但人们对它们的了解相对较少。 关于编码核心中心体成分的基因的调节，如周围素(Pcnt)样蛋白 (PLP)，一种保守的中心体支架，也是睫状体功能所必需的。完成这项提案将 加深对PLP基因转录后调控的理解。在人类中，放松对 同源的PCNT基因导致先天性疾病，如小头骨发育不良 原始矮小II型(MOPD II)和21三体。同样，在果蝇中，PLP的放松管制会导致 多种缺陷，包括胚胎致死、神经元功能障碍和男性不育。其作用机制 与PLP丢失相关的多效性表型的潜在原因尚不完全清楚。预测PLP 来编码12个mRNA变体，但是什么机制导致了这些不同的RNA物种，以及它们是如何 表达受时空调控的基因是完全未知的。在这个K99/R00提案中，我将测试 假设PLP蛋白的异构体表达，以及PLP的转录后调控 信使核糖核酸，调节其在不同组织中的不同功能。提出了三个目标来检验这一点 假设。在目标1中，我将确定胚胎PLP mRNA的定位和翻译机制。在目标2中， 我将探索替代启动子和3‘UTR用法对不同PLP RNA产生的贡献 变种。在目标3中，我将确定PLP蛋白异构体的表达谱并检测其生物学特性 包括PLPPM在内的PLP亚型在果蝇神经母细胞中的功能与早期胚胎相比。完成度 这项工作将揭示功能基因在空间和时间上不同表达模式的机制 PLP蛋白在果蝇不同组织中的异构体，将有助于我们对PLP mRNA的理解 在人类疾病中，如神经退行性疾病，其某些方面可能会被解除管制 和心血管疾病。此外，该奖项还将提供RNA方面的技术和专业培训- 测序和生物信息学、CRISPR基因组编辑和先进的成像方法 在我的专家导师的指导下。我将遵循一个有组织的培训计划，以提高我的专业能力 建立并运营我自己成功的独立实验室。

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.