Stochastic vs. Deterministic Cell Fate Choice in Fly Retinal Patterning

描述（由申请方提供）：在开发过程中，可以以两种根本不同的方式指定细胞命运。在大多数情况下，命运的决定取决于特定的信号，这些信号产生本质上是硬连线的，高度可重复的结果。在许多重要的替代情况下，分化的细胞类型通过随机机制偶然出现。第二种策略的最佳研究实例之一是果蝇视网膜中感光细胞亚型的特化。我们已经确定了一组相关的果蝇，它们以确定性和高度有序的方式做出相同的神经命运决定，产生交替的单行感光细胞亚型，而不是果蝇视网膜中随机的盐和胡椒分布。目的：我们的初步研究表明，关键转录因子Spineless的差异调节在这两个系统中是至关重要的。我们提出了一个比较的方法来表征上游和下游组件的“有序”系统，并进一步剖析这两个系统中的决定的背景和监管控制。具体目标：（1）描述交替有序系统（Dolichopodidae，或“Doli”）中的视网膜模式并测试其功能;（2）通过识别关键决定因素的调节区域，更好地理解每个细胞命运决定系统是如何控制的;（3）通过应用新开发的基因组工具，进一步剖析果蝇中感光细胞亚型的特化。研究设计：我们正在“确定性”系统（Doli）中对两个物种的基因组和转录组进行测序。这将使我们能够表征视网膜图案网络中基因的mRNA表达模式，以及关键转录因子的传统抗体开发。为了评估潜在的保守的调节功能，我们将测试候选区域从Doli在果蝇以及开发转基因技术在Doli。为了进一步评估在果蝇的命运决定的上游和下游组件的作用和功能，我们将使用FACS分选，细胞类型特异性群体的感光细胞核的转录和染色质标记分析。总体相关性：这些研究将有助于我们理解不同类型的细胞命运选择是如何控制的。两个密切相关但交替的系统的发现提供了一个理想的机会来研究潜在的生化“噪音”在关键细胞命运决定因素表达中的作用，以及如何利用或限制它。这个细胞命运规范系统的几个重要组成部分 它的靶点与癌症直接相关，包括Spineless本身的人类同源物。最终，更好地了解细胞命运是如何确定和维持的，将导致新的策略，当细胞命运被错误指定或丢失时，将治疗剂和药物靶向特定的细胞命运指定途径，这在癌症和遗传疾病的进展中经常发生。

DESCRIPTION (provided by applicant): Cell fate can be specified during development in two fundamentally different ways. In most cases, fate decisions depend on specific signals that produce essentially hardwired, highly reproducible outcomes. In a number of important alternate cases, differentiated cell types arise instead by chance, through stochastic mechanisms. One of the best studied examples of this second strategy is the specification of photoreceptor subtypes in the Drosophila retina. We have identified a related group of flies that instead make the same neural fate decision in a deterministic and highly ordered way, producing alternating single rows of photoreceptor subtypes instead of the random salt and pepper distribution in the Drosophila retina. Objectives: Our preliminary studies indicate that differential regulation of the key transcription factor Spineless is critical in both systems. We propose a comparative approach to characterize upstream and downstream components in the "ordered" system and to further dissect the context and regulatory control of this decision in both systems. Specific Aims: (1) To characterize retinal patterning in the alternate, ordered system (Dolichopodidae, or "Doli") and to test its function; (2) to better understand each how each cell fate determination system is controlled by identifying regulatory regions of critical determinants; and (3) to further dissect photoreceptor subtype specification in Drosophila by applying newly developed genomic tools. Study design: We are sequencing the genome and transcriptome of two species in the "deterministic" system (Doli). This will allow us to characterize mRNA expression patterns of genes in the retinal patterning network, alongside traditional antibody development for key transcription factors. To assess potentially conserved regulatory function we will test candidate regions from Doli in Drosophila as well as develop transgenic techniques in Doli. To further evaluate the role and function of upstream and downstream components of this fate decision in Drosophila, we will use FACS-sorted, cell-type specific populations of photoreceptor nuclei for transcriptional and chromatin mark profiling. Overall relevance: These studies will contribute to our understanding of how different types of cell fate choice are controlled. The discovery of two closely related but alternate systems provides an ideal opportunity to study the role of underlying biochemical "noise" in the expression of critical cell fate determinants and how it might be utilized - or limited. Several important components of this cell fate specification system and its targets have been implicated in cancer directly, including the human homolog of Spineless itself. Ultimately, a better understanding of how cell fate is determined and maintained will lead to new strategies for targeting therapeutic agents and drugs to specific cell fate specification pathways when cell fate is mis-specified or lost, as often occurs in cancer and in the progression of genetic diseases.