Illuminating the gene regulation underlying meiotic differentiation

摘要 减数分裂是负责配子形成的保守的分化程序。作为一个细胞 通过减数分裂分化，它经历了细胞结构和功能的单向变化 这在很大程度上是由基因表达变化驱动的。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控制减数分裂的基因调控电路 差异化。我们使用发芽酵母来研究这一过程，因为研究得很好的减数分裂因子是高度 保守的，因为这种有机体独一无二地提供了大量高度同步的细胞 这是我们常规使用的基因组方法的关键。我们的研究使蛋白质的鉴定成为可能 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控这些基因的重大惊喜。 在这些惊喜中，我们发现了一种非常规的基因调控模式，涉及到受调控的切换 在可翻译的信使核糖核酸异构体和5‘端延伸且翻译不佳的异构体之间，将被常用 随着时间的推移来驱动减数分裂蛋白质水平。我们发现这种调节模式在减数分裂中很重要，但 在其他情况下，我们研究的一个主要焦点是更好地了解它是如何工作的。虽然我们 要知道，上游开放阅读框架(UORF)负责抑制某些ORF的翻译 延伸的mRNA异构体，我们不知道为什么不是所有的情况都是这样的。我们将使用以下工具来解决此问题 记者实验，以及抑制和非抑制的mRNA结构和序列分析 成绩单。我们也不知道mrna的降解如何影响这一调控和减数分裂基因。 更广泛的表达，我们将使用新的代谢标记方法进行研究。 除了对已知基因的非常规调控外，我们还发现减数分裂细胞翻译 许多基因之前都没有被识别出来。其中包括数百个基因，它们以 非8月密码子，以及比用于注释的100密码子截止值短的数千个密码子 基因组。我们已经验证了这些非规范蛋白的表达，现在正在研究 它们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翻译启动在减数分裂中很常见，主要使用对候选调控因子的研究 我们已经确定了。我们正在进行联合筛选，以确定许多短减数分裂蛋白的作用，以及 对短蛋白包括特征蛋白结构域的情况进行定向研究。团结在一起 这里提出的项目将解释减数分裂细胞如何以及为什么使用非规范的基因调控。 特征，我们认为这是解开减数分裂进程的分子控制的关键。

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.