Functional genomic analysis of the haploinsufficient tumor suppressor, CUX1

CUX1 is a homeodomain-containing transcription factor that is conserved, ubiquitous, and essential in vertebrates and invertebrates. CUX1 is mutated or deleted in high-risk myeloid neoplasms and solid tumors, resulting in haploinsufficiency and tumor growth. Here we provide the first analysis of endogenous, whole genome, CUX1 DNA binding. We demonstrate that CUX1 binds with transcriptional activators and cohesin at distal enhancers across three different human cell types. Haploinsufficiency of CUX1 altered the expression of a large number of genes, including cell cycle regulators, with concomitant increased cellular proliferation. Surprisingly, CUX1 occupancy decreased genome-wide in the haploinsufficient state, and binding site affinity did not correlate with differential gene expression. Instead, differentially expressed genes had multiple, low-affinity CUX1 binding sites, consistent with an analog model for CUX1 gene regulation. A machine-learning algorithm determined that chromatin accessibility, enhancer activity, and distance to the transcription start site are features of functional CUX1 DNA binding. Moreover, CUX1 is enriched at sites of DNA looping, and these loops connect CUX1 to the promoters of target genes. We propose that CUX1 is an analog transcription factor that regulates target genes through higher order genome architecture. RNA-seq and ChIP-seq of CUX1 in the wildtype and haploinsufficient states in K562 cells

CUX1是一种含同源异型结构域（homeodomain）的转录因子，在脊椎动物与无脊椎动物中均具有保守性、普遍存在且不可或缺。CUX1在高危髓系肿瘤及实体瘤中发生突变或缺失，会引发单倍体剂量不足（haploinsufficiency）并促进肿瘤生长。本研究首次针对内源性全基因组CUX1的DNA结合情况展开分析。我们证实，在三种不同的人类细胞类型中，CUX1可与转录激活因子及黏连蛋白（cohesin）结合于远端增强子区域。CUX1单倍体剂量不足会改变大量基因的表达，其中包括细胞周期调控因子，并伴随细胞增殖能力的显著增强。令人意外的是，在单倍体剂量不足状态下，全基因组范围内CUX1的结合占有率整体下降，且结合位点的亲和力与差异基因表达并无关联。与之相反，差异表达基因带有多个低亲和力的CUX1结合位点，这与CUX1基因调控的类比模型相符。机器学习算法确定，染色质可及性、增强子活性以及与转录起始位点的距离，是决定CUX1 DNA结合功能的核心特征。此外，CUX1在DNA环化位点处富集，且此类环化结构可将CUX1与其靶基因的启动子相连。我们提出，CUX1属于一类通过高阶基因组架构调控靶基因的类比转录因子。本数据集包含K562细胞在野生型与单倍体剂量不足状态下的CUX1 RNA测序（RNA-seq）及染色质免疫共沉淀测序（ChIP-seq）数据。