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

总结 像眼睛这样复杂器官的发育涉及多种不同器官的特化。 细胞类型及其功能的整合。在果蝇中，光感受器和 分泌透镜并产生屏蔽色素的神经胶质样视锥细胞和色素细胞来自单个 等价的祖先的领域。这些细胞命运之一的承诺需要细胞外信号， 与内在转录因子的复杂网络整合。尽管受体酪氨酸激酶 多年来已知信号传导诱导光感受器和视锥细胞，其大多数靶细胞是光感受器和视锥细胞。 基因仍然未知。也不清楚哪些内在转录因子区分这些细胞身份。 锌指转录因子Glass曾被认为是决定感光细胞命运的因子，但我们最近的研究发现， 研究表明，它在三种细胞类型中的每一种中都起作用，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。第一个目标将 寻找视网膜分化过程中受体酪氨酸激酶信号转导和Glass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 Pointed是一种介导表皮生长因子受体（EGFR）信号传导的转录因子， 未分化细胞、光感受器和视锥细胞。类似的方法将用于识别玻璃目标 基因，这两个数据集将与改变其表达的基因的转录组学分析进行比较 在Egfr或玻璃突变体。这些实验的目的是确定转录因子， 通过EGFR信号传导促进光感受器和视锥细胞的分化，并阐明玻璃 加入到他们的规则中第二个目标是关于转录抑制因子如何限制转录因子的作用。 玻璃驱动细胞类型特异性基因表达。两个明确的调控区域， 以玻璃依赖性方式的光受体特异性表达将用于鉴定 阻止玻璃激活这些基因在锥细胞和色素细胞。这些抑制因子的重要性 结合位点将在内源基因组环境中进行测试。在补充方法中，增强剂 将鉴定在视锥细胞或色素细胞中特异性驱动玻璃依赖性表达的区域， 其特点是规则。结合起来，这些实验将揭示不同的细胞身份如何能够 使用有限的一组信号和转录因子从共同的祖细胞中指定，这一过程 也发生在哺乳动物的视网膜中。这些结果将有助于完善我们生产特定细胞类型的能力 用于再生医学。

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.