一种基因组来源的动态癌细胞状态非编码报告子

Sequential changes in transcriptional cell state are essential for normal development and are coopted in cancer. Controlling these changes for therapeutic benefit has been limited due to a lack of tools that reflect different states in live cells. Here, we describe a new reporter method termed “TRECS” that integrates epigenomic and transcriptomic measurements to define endogenous genomic elements that label individual cells in different cell states. We use TRECS in the neural crest-derived, high-risk pediatric solid tumor neuroblastoma, where we demonstrate broad presence of cells with distinct transcriptomes, associated with functional chemoresistance and sensitivity. Experiments using neural crest stem cells and TRECS mouse knock-in model identified that TRECS-labelled cells reflect early developmental stages in the neural crest. These cells display real-time plasticity of transcriptional state and phenotype, in a manner unlinked to cell cycle control. Investigation of nominated loci demonstrates state-specific enrichment of elements marked by H3K27ac, H3K4me1 and open chromatin by ATAC-seq, which flexibly change as cells transition between these phenotypically divergent states. To investigate whether the primary nominated locus is a driver or reporter of cell state, we integrate micro-C, transcriptomics, truncation experiments and functional CRISPRi to identify that this region functions as a pure endogenous reporter of cell state. This, therefore, provides a mechanism to identify new, state-controlling transcription factors. Motif analysis demonstrated enrichment of AP1 transcription factor motifs in the chemoresistant state, and knockout of these AP1 transcription factors results in rewiring of cell state and enhanced chemosensitivity without effects on cell growth. To capitalize on the endogenous flexibility in this system and identify mechanisms to enforce cell state changes independent of cell growth and death, we performed high-content image-based small molecule screening to identify targets suitable to enhance chemosensitivity. These experiments identified EP300/CBP, master histone acetyltransferases, as crucial controllers of a primitive, chemoresistant cell state. Transient acetyltransferase and bromodomain-based inhibition of EP300/CBP results in transcriptional and epigenetic reprogramming in vitro and in vivo , leading to enhanced chemosensitivity and prolonged survival in murine models. These results demonstrate an unbiased method to identify non-coding genomic loci enriched in specific cell states, which can be harnessed to identify master transcription factors driving these cell states and similarly, mechanisms to enforce changes in cell state.