Integration of Diverse Inputs Determines Developmental Outcomes

 描述(由申请人提供)：本项目的重点是阐明噬菌体Lambda感染后在大肠杆菌中裂解/溶源决策的潜在机制，这是一个决定细胞命运和发育遗传网络的范例系统。这一系统提供了极具吸引力的机会来研究单个基因表达的随机性和过程中涉及的空间方面如何有助于涉及调控遗传网络的决策过程的随机性-决定论二元性。通过我们最近使用活细胞显微镜进行的研究，我们假设了一个与经典图景不同的令人惊讶的决策场景。然而，潜在的机制仍不清楚。我们建议在单细胞/单病毒/单分子水平上对该系统进行更详细的系统研究。该提案有三个具体目标。第一个目标集中在阐明个体噬菌体基因组如何在裂解/溶源决定中起作用。它包括探索细胞内噬菌体DNA的时空信息如何与细胞命运相关，在单噬菌体DNA水平上如何改进对已知确定性因素(如每细胞感染噬菌体的数量和细胞大小)的裂解/溶源决策的描述，以及通过混合感染实验噬菌体如何相互合作或竞争。第二个目标是系统地确定裂解/溶源对不同生长条件和噬菌体因子的敏感性和响应性，并用高分辨率和高通量的方法研究基因表达。最终目的是建立考虑噬菌体复制的基因表达的数学和计算模型，解释各种遗传和环境因素对裂解/裂解决定的影响。总体而言，我们希望对各种确定性因素和随机性如何有助于一个众所周知但尚未完全理解的发展过程的结果达成定量的理解。

 DESCRIPTION (provided by applicant): The focus of this project is to elucidate the underlying mechanism for lysis/lysogeny decision making in E. coli upon infection by phage lambda, a paradigmatic system for cell-fate determination and developmental genetic networks. This system offers appealing opportunities to examine how the stochasticity of gene expression from a single gene and spatial aspects involved in the process contribute to the stochasticity-determinism duality nature of the decision-making process which involves a regulatory genetic network. Through our recent study using live-cell microscopy, we hypothesized a different and surprising decision-making scenario from the classical picture. However, the underlying mechanism remains unclear. We propose a systematic investigation of this system in much more detail at the single-cell/single-virus/single-molecule level. The proposal has three specific aims. The first Aim focuses on elucidating how individual phage genome contributes to the lysis/lysogeny decision. It includes exploring how the spatiotemporal information of phage DNA inside the cell correlates with cell fates, how the description of lysis/lysogeny decision in response to the known deterministic factors (e.g. number of infecting phages per cell and cell size) is improved when examined at the single phage-DNA level, and how phages cooperate or compete with each other through mixed infection experiments. The second Aim focuses on determining the sensitivity and responsiveness of the lysis/lysogeny over a full spectrum of distinct growth conditions and phage factors systematically and investigating the gene expression with high-resolution and high-throughput methods. The final Aim is to develop mathematical and computational models of gene expression that account for phage replication, to explain the influence of various genetic and environmental factors on the lysis/lysogeny decision. Overall, our hope is to reach a quantitative understanding of how various deterministic factors and stochasticity contribute to the outcome of a well-known, but incompletely understood developmental process.