MFB: Massively parallel identification of translation regulatory sequences in human and viral mRNAs

信使RNA（mRNA）编码制造蛋白质的指令，蛋白质构成细胞功能的基本机制。最近的技术进步使得能够开发出可以递送给人类的治疗性mRNA，特别是在广泛使用的SARS-CoV-2疫苗中。这些疗法的成功表明了新一代mRNA药物的潜力，其应用范围超出了疫苗，如抗癌疗法和遗传疾病的治疗。实现这些目标将需要设计优化蛋白质表达的mRNA，并且可以针对特定组织和细胞环境进行定制。该项目的目标是利用一种策略来量化数千种RNA的合成文库的翻译功能，以发现改变蛋白质产生的时间和数量的特征。深入了解基因表达的这些基本规则将是设计新的mRNA治疗方法的重要基石，以解决更广泛的人类疾病，从而推进RNA生物技术。该项目还将为博士后学者提供培训机会，并吸引初中和高中学生参与RNA生物学。由mRNA合成的蛋白质数量可以跨越两个数量级以上，在不同的细胞类型中变化，并根据细胞信号迅速变化。决定这一表达范围的mRNA的特征在很大程度上仍然未知。可能包括RNA结合蛋白识别的序列基序、结构化RNA元件和影响翻译过程的核苷酸修饰。该项目将采用最近开发的大规模平行报告基因检测方法，系统地鉴定数千个人类和病毒5′ UTR的特征，这些特征决定了蛋白质产生的数量和时间。目标1将检查不同类型的核苷酸修饰的翻译功能，并确定识别它们的分子机制。目标2将定量从环状RNA的合成文库中产生的蛋白质，以鉴定能够进入非经典翻译起始机制的RNA元件。目的3将确定确定的mRNA的功能，确定细胞类型的特异性表达和建立这种特异性的机制。这些努力将提供对mRNA编码的翻译决定因素的全面理解，提供对基因表达的基本决定因素的见解，并指导新的mRNA疗法的设计。该项目由遗传机制计划/生物科学理事会分子和细胞生物科学部以及数学和物理科学理事会化学部。该奖项反映了NSF的法定使命，并已被认为是值得通过使用基金会的知识价值和更广泛的影响审查标准进行评估的支持。

Messenger RNAs (mRNAs) encode instructions for making proteins, which constitute the fundamental machinery for cellular function. Recent technological advancements have enabled the development of therapeutic mRNAs that can be delivered to humans, notably evident in widely used vaccines for SARS-CoV-2. The success of these therapeutics suggests the potential of new generations of mRNA medicines with applications beyond vaccines, such as anti- cancer therapies and treatment for genetic disorders. Realizing these goals will require the design of mRNAs that optimize protein expression and can be customized for specific tissues and cellular environments. The goal of this project is to leverage a strategy for quantifying the translation functions of synthetic libraries of thousands of RNAs to discover features that modify the timing and quantity of protein production. Insights into these fundamental rules for gene expression will be important building blocks for engineering new classes of mRNA therapeutics to address a broader spectrum of human disease, thus advancing RNA biotechnology. The project will also provide training opportunities for postdoctoral scholars and engage middle and high school students in RNA biology.The quantity of protein synthesized from an mRNA can span more than two orders of magnitude, vary across cell types, and rapidly change in response to cellular signals. The features of mRNAs that determine this range of expression remain largely unknown. Possibilities include sequence motifs recognized by RNA-binding proteins, structured RNA elements and nucleotide modifications that affect the translation process. This project will employ a recently developed massively parallel reporter assay to systematically identify features of thousands of human and viral 5′ UTRs that determine the amount and timing of protein production. Aim 1 will examine the translation functions of diverse classes of nucleotide modifications and identify the molecular mechanisms that recognize them. Aim 2 will quantify protein production from a synthetic library of circular RNAs to identify RNA elements capable of accessing mechanisms for non-canonical translation initiation. Aim 3 will identify features of mRNAs that determine cell-type specific expression and the mechanisms that establish this specificity. These efforts will provide a comprehensive understanding of mRNA- encoded determinants of translation, providing insights into basic determinants of gene expression and guiding the design of new mRNA therapeutics.This project is supported by the Genetic Mechanisms program/Division of Molecular and Cellular Biosciences in the Directorate for Biological Sciences and by the Division of Chemistry in the Directorate for Mathematical and Physical Sciences.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.