Diversification of cell types in the Drosophila retina - Resubmission - 1

摘要 像眼睛这样的复杂器官的发育涉及到多个不同的 细胞类型及其功能的整合。在果蝇中，光探测光感受器和 分泌晶状体并产生筛选色素的神经胶质样锥体和色素细胞来自于单个 相当于祖先的领域。对这些细胞命运之一的承诺需要细胞外信号 整合了一个复杂的内在转录因子网络。尽管受体酪氨酸激酶 多年来，人们已经知道信号既可以诱导光感受器，也可以诱导视锥细胞，这是它的主要靶点 基因仍然未知。目前也不清楚哪些内在转录因子区分了这些细胞身份。 锌指转录因子Glass曾被认为决定了光感受器的命运，但我们最近 研究表明，它作用于这三种类型的细胞中的每一种，以促进它们的正常分化。这项建议 试图了解这些共同因素如何激活不同的分化途径。第一个目标是 寻找受体酪氨酸激酶信号和玻璃在视网膜分化过程中的直接靶基因。 靶向DNA腺嘌呤甲基转移酶鉴定(DamID)将用于鉴定直接靶基因 介导表皮生长因子受体(EGFR)信号转导的转录因子，在 未分化细胞、光感受器和视锥细胞。将使用类似的方法来识别Glass目标 基因，这两个数据集将与改变其表达的基因的转录分析进行比较 在EGFR或玻璃突变体中。这些实验的目标是识别被诱导的转录因子 通过EGFR信号促进光感受器和视锥细胞的分化，并阐明玻璃是如何 会影响到他们的监管。第二个目的是关于转录抑制因子如何限制 玻璃来驱动特定细胞类型的基因表达。两个已定义的监管区域，已知推动 玻璃依赖方式的光感受器特异性表达将被用来识别 防止玻璃激活锥体细胞和色素细胞中的这些基因。这些抑制者的重要性 结合位点将在内源基因组环境中进行测试。在一个补充方法中，增强器 将确定在视锥细胞或色素细胞中驱动玻璃依赖表达的区域，并 他们的规定具有鲜明的特点。结合起来，这些实验将揭示不同的细胞身份如何 使用一组有限的信号和转录因子从共同的祖细胞中指定，这一过程 哺乳动物的视网膜中也存在这种情况。结果将有助于提高我们产生特定细胞类型的能力 用于再生医学的干细胞。

Summary The development of a complex organ such as the eye involves the specification of multiple distinct cell types and the integration of their functions. In Drosophila, the light-detecting photoreceptors and the glial-like cone and pigment cells that secrete the lens and produce screening pigments arise from a single field of equivalent progenitors. Commitment to one of these cell fates requires extracellular signals to be integrated with a complex network of intrinsic transcription factors. Although receptor tyrosine kinase signaling has been known for many years to induce both photoreceptors and cone cells, most of its target genes remain unknown. It is also unclear which intrinsic transcription factors distinguish these cell identities. The zinc finger transcription factor Glass had been thought to specify the photoreceptor fate, but our recent work showed that it acts in each of the three cell types to promote their normal differentiation. This proposal seeks to understand how these common factors activate distinct differentiation pathways. The first aim will look for direct target genes of receptor tyrosine kinase signaling and Glass during retinal differentiation. Targeted DNA adenine methyltransferase identification (DamID) will be used to identify direct target genes of Pointed, the transcription factor that mediates Epidermal growth factor receptor (EGFR) signaling, in undifferentiated cells, photoreceptors and cone cells. A similar approach will be used to identify Glass target genes, and both datasets will be compared to transcriptomic analysis of genes that change their expression in Egfr or glass mutants. The goal of these experiments is to identify transcription factors that are induced by EGFR signaling to promote the differentiation of photoreceptors and cone cells, and elucidate how Glass feeds into their regulation. The second aim concerns how transcriptional repressors restrict the effects of Glass to drive cell type-specific gene expression. Two defined regulatory regions that are known to drive photoreceptor-specific expression in a Glass-dependent manner will be used to identify repressors that prevent Glass from activating these genes in cone and pigment cells. The importance of these repressor binding sites will be tested in the endogenous genomic context. In a complementary approach, enhancer regions that drive Glass-dependent expression specifically in cone or pigment cells will be identified and their regulation characterized. In combination, these experiments will reveal how distinct cell identities can be specified from common progenitors using a limited set of signals and transcription factors, a process that also occurs in the mammalian retina. The results will help to refine our ability to produce specific cell types from stem cells for regenerative medicine.