Illuminating the gene regulation underlying meiotic differentiation

阐明减数分裂分化背后的基因调控

• 批准号: 10545416
• 负责人: Gloria Ann Brar
• 金额: $ 8.3万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-01-01 至 2024-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10545416
• 关键词: Address; Case Study; Cell physiology; Cells; Cellular Structures; Codon Nucleotides; Event; Gene Expression; Gene Expression Regulation; Genes; Genome; Genomic approach; Germ Cells; Label; Meiosis; Messenger RNA; Metabolic; Modeling; Molecular; Open Reading Frames; Organism; Process; Production; Protein Isoforms; Proteins; Regulation; Regulator Genes; Reporter; Research; Role; Saccharomycetales; Sexual Reproduction; Structure; Time; Transcript; Translating; Translation Initiation; Translations; Work; base; expectation; experimental study; mRNA Transcript Degradation; precursor cell; programs

ABSTRACT Meiosis is the conserved differentiation program that is responsible for gamete formation. As a cell progress through meiotic differentiation, it undergoes unidirectional changes in cellular structure and function that are largely driven by gene expression changes. Because the molecular basis for most meiotic transitions remains mysterious, my lab aims to illuminate the gene regulatory circuitry that programs meiotic differentiation. We use budding yeast to study this process because well studied meiotic factors are highly conserved and because this organism uniquely offers access to the large number of highly synchronous cells that is key to genomic approaches that we routinely employ. Our studies have enabled identification of proteins involved in key meiotic processes, and new regulatory events during meiosis. These studies have also uncovered major surprises in the genes that meiotic cells express and how they regulate these genes. Among these surprises, we found an unconventional mode of gene regulation, involving regulated toggling between a translatable mRNA isoform and one that is 5’ extended and poorly translated, to be commonly used to drive meiotic protein levels over time. We have found this mode of regulation to be important in meiosis but also in other conditions, and a major focus of our research is to better understand how it works. Although we know that upstream open reading frames (uORFs) are responsible for repressed ORF translation on some extended mRNA isoforms, we do not know why this is not true of all cases. We will address this question using reporter experiments, and analysis of mRNA structures and sequences of repressed versus non-repressed transcripts. We also do not understand how mRNA degradation impacts this regulation and meiotic gene expression more broadly, which we will study using new metabolic labeling approaches. Beyond unconventional regulation of known genes, we also discovered that meiotic cells translate many genes were not previously identified. These include hundreds of genes that are translated starting with non-AUG codons, and thousands that are shorter than the 100 codon cutoff that was used to annotate genomes. We have validated the expression of these non-canonical proteins and are now studying the molecular mechanisms underlying their synthesis and their specific cellular roles. We are investigating why non-AUG translation initiation is common in meiosis, primarily using study of candidate regulatory factors that we have identified. We are performing pooled screens to identify roles for the many short meiotic proteins, and directed study of cases in which the short proteins include domains of characterized proteins. Together the projects proposed here will explain how and why meiotic cells employ non-canonical gene regulatory features, which we believe is critical to unraveling the molecular control of meiotic progression.

摘要 减数分裂是负责配子形成的保守的分化程序。作为细胞 通过减数分裂分化，它经历了细胞结构和功能的单向变化 主要由基因表达变化驱动。因为大多数减数分裂转变的分子基础 仍然神秘，我的实验室旨在阐明基因调控电路程序减数分裂 分化我们使用芽殖酵母来研究这一过程，因为研究得很好的减数分裂因素是高度 保守，因为这种生物体独特地提供了大量的高度同步细胞， 这是我们常规使用的基因组方法的关键。我们的研究使我们能够鉴定 参与关键的减数分裂过程，以及减数分裂过程中的新调控事件。这些研究还 发现了减数分裂细胞表达的基因以及它们如何调节这些基因的重大惊喜。 在这些惊喜中，我们发现了一种非传统的基因调控模式，涉及到调控切换 在可翻译的mRNA同种型和5'延伸且翻译差的mRNA同种型之间，通常使用 来驱动减数分裂的蛋白质水平。我们发现这种调节方式在减数分裂中很重要， 我们研究的一个主要重点是更好地了解它是如何工作的。虽然我们 我们知道上游开放阅读框架（uORF）负责一些 延伸的mRNA亚型，我们不知道为什么这不是所有情况下都是如此。我们将使用 报告实验，并分析mRNA结构和序列的抑制与非抑制 成绩单我们也不知道mRNA降解如何影响这种调控和减数分裂基因 表达更广泛，我们将使用新的代谢标记方法研究。 除了已知基因的非常规调控，我们还发现减数分裂细胞翻译 许多基因以前没有被鉴定。其中包括数百个基因，这些基因以 非AUG密码子，以及数千个短于100密码子截止值的密码子， 基因组我们已经验证了这些非典型蛋白质的表达，现在正在研究这些蛋白质的表达。 其合成的分子机制及其特定的细胞作用。我们正在调查原因 非AUG翻译起始在减数分裂中是常见的，主要使用候选调节因子的研究， 我们已经确定。我们正在进行合并筛选，以确定许多短减数分裂蛋白的作用， 对短蛋白质包括特征蛋白质的结构域的情况进行定向研究。一起 这里提出的项目将解释减数分裂细胞如何以及为什么采用非经典基因调控 特征，我们认为这是解开减数分裂进程的分子控制的关键。