SINGLE-CELL CHEMICAL TRANSCRIPTOMIC DISSECTION OF AN ESSENTIAL TRANSCRIPTION FACTOR NETWORK

一种必需转录的单细胞化学转录切割 要素网络 项目摘要/摘要 延伸的MYC网络中的碱性螺旋-环-螺旋亮氨酸拉链转录因子(TF)起着至关重要的作用 调节细胞生长、分化和动态平衡。开创性研究表明MYC及其 在发育和癌症中作为增殖和新陈代谢驱动力的转录因子的相互作用网络，但如何剂量 在这些因素中，转录状态的编码仍然存在争议。在这里，我提出了一种新的化学基因组 系统地确定转铁蛋白的剂量、转录产量和细胞状态是如何联系起来的。这就做 将我们最近发表的大规模多重单细胞RNA-SEQ筛选方法(SCI-Plex)与 蛋白质的化学遗传降解以研究胚胎干细胞是如何转录的 在TF的剂量降解后重新编程。然后，我将使用最先进的计算工具来 量化由特定剂量编码的细胞状态轨迹。最后，我将利用这个单细胞 《微扰图谱》支持基因组图谱实验，以了解如何减少这些因素的剂量 在体内导致整个基因组的物理重新分布。这些研究将用于。)阐明以下机制： Tf剂量编码转录输出，ii。)阐明相互作用的因子在细胞内的浓度 保持染色质、转录和细胞状态，以及提供机械的洞察力，了解 转录因子网络与表观基因组相互作用，调节哺乳动物的多能性。更广泛地说， 这些方法有可能解决长期存在的基因调控问题，如蛋白质剂量 控制健康和癌症中的细胞状态。

SINGLE-CELL CHEMICAL TRANSCRIPTOMIC DISSECTION OF AN ESSENTIAL TRANSCRIPTION FACTOR NETWORK PROJECT SUMMARY / ABSTRACT Basic helix-loop-helix leucine zipper transcription factors (TFs) in the extended MYC network play essential roles regulating cellular growth, differentiation, and homeostasis. Seminal studies have implicated MYC and its interacting network of TFs as drivers of proliferation and metabolism in development and cancer, but how dosage of these factors encodes transcriptional state remains contentious. Here, I propose a novel chemical genomic framework to systematically determine how TF dosage, transcriptional output, and cellular state are linked. I will combine our recently published massively multiplex single-cell RNA-seq screening method (sci-Plex) with chemical genetic degradation of protein targets to examine how embryonic stem cells are transcriptionally reprogrammed following dosed degradation of TFs. I will then employ state-of-the-art computational tools to quantify the cellular state trajectories encoded by specific dosages. Finally, I will leverage this single-cell ‘perturbation atlas’ to support genomic mapping experiments to understand how reduced dosage of these factors in vivo leads to physical redistribution across the genome. These studies will i.) illuminate mechanisms by which TF dosages encode transcriptional output, ii.) elucidate how intracellular concentrations of interacting TFs maintain chromatin, transcriptional, and cellular states, and iii.) provide mechanistic insight into how a transcription factor network interacts with the epigenome to regulate mammalian pluripotency. More generally, these approaches have the potential to address long-standing gene regulatory questions of how protein dosage controls cellular state in both health and in cancer.