Developmental priming of mRNA decay during Drosophila embryogenesis

摘要 销毁储存的母体转录本是动物早期胚胎发育过程中必不可少的一步。 与产生新的受精卵转录本一起，这个过程被称为母体到受精卵的转变 (MZT)，并使发育控制从母体基因组移交给合子基因组。 信使核糖核酸的破坏是由3‘聚(A)尾的缩短触发的，尾长的控制是一个重要的因素 转录后调控中心。然而，我们对Poly(A)-尾部长度的差异知之甚少 母体和受精卵的成绩单。我们提出了一个“发展启动”假说来弥合这一概念。 GAP：在这里，母体转录经历了卵子发生特有的Poly(A)尾巴缩短，这导致了他们的 在MZT期间的毁灭。因此，发育启动提供了一种机制，即母体和 合子转录本可以被区分，以使发育控制转移到合子基因组。 在这项提案中，我们将以果蝇作为模型系统来检验和探索这一假说。我们会 首先关注一个潜在的发育启动原型：母性致死(SXL)mRNA。SXL是一种 性别决定和剂量补偿的高级调节器。性染色体数量导致不同的 男性和女性SXL转录异构体，而女性异构体启动了一个自动调节的前馈循环 这最终导致了女性身份的维护。重要的是，母体(以及女性)的SXL mRNA 沉积在胚胎中，必须将其移除，以免超过受精卵的性染色体组成 并导致所有胚胎都被指定为“雌性”。然而，其分子机制尚不清楚。我们 认为母系SXL基因的去除可以用发育启动来解释。在目标1中，我们 将探索这一模型：在卵子发生过程中，RNA结合蛋白Bruno导致短的Poly(A)尾巴 母体的SXL mRNA，因此使它们处于破坏状态。在目标2中，我们将扩展我们的假设转录组- Wide使用了一种新的方法，将基于SNP的等位基因检测与长阅读测序相结合来确定 母体和受精卵转录本上的多聚(A)尾巴长度。第二个目标是探索性的，非常适合。 R21机制。 我们希望我们提出的研究将做出几个重大贡献。首先，它将提供一个 解答了数十年来关于SXL在胚胎发生过程中的调控的谜团。第二，通过我们的 在目标2的实验中，我们将创建跨越果蝇的高质量长阅读测序数据集 MZT。第三，我们将开发一种新的实验和计算工具，可以使用长阅读测序 来看看等位基因的特定调控，这将使未来的机械性实验成为可能。最后，它将提供一个 理解产妇成绩单的具体清除的新概念框架，这将形成 为将来的学习和资助奠定基础。这一智力框架可能会被证明对理解 在其他发育转变时，多长时间的成绩单可以被具体调控。

SUMMARY Destruction of deposited maternal transcripts is an essential step during early animal embryogenesis. Together with production of new zygotic transcripts, this process is called the maternal-to-zygotic transition (MZT) and enables the handover of developmental control from the maternal genome to the zygotic one. mRNA destruction is triggered by shortening of the 3' poly(A) tail, and control of tail length is an important post-transcriptional regulatory hub. However, we know little about the differences in poly(A)-tail lengths on maternal and zygotic transcripts. We propose a “developmental priming” hypothesis to bridge this conceptual gap: here, maternal transcripts experience oogenesis-specific poly(A)-tail shortening that leads to their destruction during the MZT. Developmental priming thus provides a mechanism whereby maternal and zygotic transcripts can be distinguished to enable the transfer of developmental control to the zygotic genome. In this proposal, we will test and explore this hypothesis using Drosophila as a model system. We will first focus on a potential archetype of developmental priming: the maternal Sex lethal (Sxl) mRNA. Sxl is a master regulator of sex determination and dosage compensation. Sex chromosome number leads to different male and female Sxl transcript isoforms, and the female isoform sets off an auto-regulatory, feed-forward loop that culminates in the maintenance of female identity. Importantly, the maternal (and thus female) Sxl mRNA is deposited in the embryo and must be removed lest it override the zygotic sex chromosome composition and lead to all embryos being specified as “female.” However, the molecular mechanism is unknown. We propose that the removal of maternal Sxl mRNA can be explained by developmental priming. In Aim 1, we will explore this model: during oogenesis the RNA-binding protein Bruno leads to short poly(A) tails on maternal Sxl mRNA and so primes them for destruction. In Aim 2, we will extend our hypothesis transcriptome- wide using a new approach that combines SNP-based allele detection with long-read sequencing to determine the poly(A)-tail lengths on maternal and zygotic transcripts. This second aim is exploratory and perfectly suited to the R21 mechanism. We expect our proposed research to make several significant contributions. First, it will provide an answer to a decades-old mystery about Sxl regulation during embryogenesis. Second, through our experiments in Aim 2, we will create high-quality long-read sequencing datasets spanning the Drosophila MZT. Third, we will develop a new experimental and computational tool that can use long-read sequencing to look at allele-specific regulation, which will enable future mechanistic experiments. Finally, it will provide a new conceptual framework for understanding the specific clearance of maternal transcripts, which will form the basis for future studies and grants. This intellectual framework will likely prove useful for understanding how old transcripts can be specifically regulated at other developmental transitions.