Actin cytoskeleton from nucleus to organism

项目摘要/摘要 教科书告诉我们，肌动蛋白细丝赋予细胞它们的形状，并由一系列蛋白质驱动 当细胞改变形状时，肌动蛋白重塑。但在这种简单的讲述中，缺少的是一种整体的理解 上游基因表达和信号如何控制肌动蛋白重塑，不同蛋白质如何协同工作 为了重塑肌动蛋白，以及下游细胞形状的变化如何转化为及时和可靠的生物输出- 来了。由于基于肌动蛋白的失败可能源于重建之前、期间和之后的事件，我们需要一个 综合理解肌动蛋白在健康和疾病中的关键作用。 为了获得肌动蛋白的这种全景图，我的实验室研究了细胞化，这是第一个组织构建 果蝇胚胎中的事件。我们开发了这个简单的实验系统，这样我们就可以研究肌动蛋白 推动细胞化的重塑，同时也将重塑与基因水平上的上游事件联系起来 表达和信号，以及下游结果，包括形态发生保真度和胚胎存活率。 我们的方法结合了果蝇遗传学和胚胎学与定量活细胞成像的mRNA，肌动蛋白， 和肌动蛋白调节蛋白，精确到单分子分辨率。 我们的长期目标是了解肌动蛋白细胞骨架如何与亚细胞过程相互作用。 (如转录)和系统(如细胞核)，以“正确”的动力学协调细胞形状的变化，稳健- Ness和机械特性，以实现成功的生物结果。未来五年，我们将重点 根据我们正在进行的研究产生的三个目标：目标1.确定基因表达如何调节肌动蛋白重塑。 ELING-基因表达指导形态发生。然而，我们还不知道转录动力学是如何告知 肌动蛋白重塑。对于细胞化，必须转录五个编码肌动蛋白调节器的基因。我们将测试一个 认为这些基因转录的数量特征支持全球同步性和一致性的假设 胚胎中的细胞化。目标2.确定肌动球蛋白收缩的机制-肌动球蛋白收缩 对细胞形状的改变是必不可少的，但其机制存在争议。在细胞化过程中，肌动球蛋白环 在机械上不同的背靠背阶段收缩(肌球蛋白-2依赖与独立)。我们 将决定肌动蛋白结合蛋白如何驱动每种机制。目标3.确定肌动蛋白细胞骨架如何 对环境应激的反应-肌动蛋白被越来越多地认为是应激反应的中介。我们再- 最近在胚胎中发现了一种热诱导肌动蛋白应激反应(ASR)。我们将检验ASR的假设 通过改变细胞质和细胞核中的游离肌动蛋白池之间的动态平衡，使胚胎的存活率处于危险之中。 这些目标是相互促进的，因此我们将了解在实现这些目标之前、期间和之后的机制 肌动蛋白重塑共同决定胚胎的结局。我们的努力得到了我的实验室的帮助 经过验证的能力，能够量化表型和跨尺度和亚细胞系统的相关事件。这些蛋白质和 我们研究的过程在有机体中是保守的，因此我们的发现将具有广泛的相关性。

PROJECT SUMMARY/ABSTRACT Textbooks teach us that actin filaments give cells their shape, and that a “parts list” of proteins drives actin remodeling when cells change shape. But what is missing from this simple telling is a holistic understanding of how upstream gene expression and signaling control actin remodeling, how different proteins work together to remodel actin, and how downstream cell shape change is converted into timely and reliable organismal out- comes. Because actin-based failures can stem from events before, during and after remodeling, we need an integrated understanding to make sense of actin’s critical role in health and disease. To obtain this kind of “whole picture” view of actin, my lab studies cellularization, the first tissue-building event in Drosophila embryos. We developed this simple experimental system so that we can study the actin remodeling that drives cellularization, while also relating that remodeling to upstream events at the level of gene expression and signaling, and downstream outcomes including morphogenetic fidelity and embryonic viability. Our methods combine Drosophila genetics and embryology with quantitative live-cell imaging of mRNAs, actin, and actin regulatory proteins, down to single-molecule resolution. Our long-term objective is to understand how the actin cytoskeleton interacts with subcellular processes (e.g. transcription) and systems (e.g. nucleus) to orchestrate cell shape change with “the right” kinetics, robust- ness and mechanical properties to achieve successful organismal outcomes. In the next five years, we will focus on three goals arising from our ongoing studies: Goal 1. Determine how gene expression regulates actin remod- eling – Gene expression instructs morphogenesis. Yet, we do not know how transcriptional dynamics inform actin remodeling. For cellularization, five genes that encode actin regulators must be transcribed. We will test a hypothesis that quantitative features of transcription of these genes underpin the global synchrony and uniformity of cellularization in embryos. Goal 2. Determine mechanisms of actomyosin contraction – Actomyosin contraction is essential to cell shape change, but its mechanism is controversial. During cellularization, actomyosin rings contract in back-to-back phases that are mechanistically distinct (Myosin-2 dependent versus independent). We will determine how actin binding proteins drive each mechanism. Goal 3. Determine how the actin cytoskeleton responds to environmental stress – Actin is increasingly recognized as a mediator of stress response. We re- cently identified a heat inducible Actin Stress Response (ASR) in embryos. We will test the hypothesis that ASR puts embryo viability at risk by altering homeostasis between free actin pools in the cytoplasm and nucleus. These goals build on each other so that we will understand how mechanisms before, during and after actin remodeling work together to determine outcomes for the embryo. Our efforts are facilitated by my lab’s proven ability to quantify phenotypes and relate events across scales and subcellular systems. The proteins and processes we study are conserved across organisms so our findings will be broadly relevant.