ATAC-seq analysis of K562, MKPL-1, MV4-11, NB4 and U937 leukemia cell lines. ATAC-seq analysis of K562, MKPL-1, MV4-11, NB4 and U937 leukemia cell lines

Mutations in protein-coding genes are well established as the basis for human cancer, yet it remains elusive how alterations within non-coding genome, a substantial fraction of which contain cis-regulatory elements (CREs), contribute to cancer pathophysiology. Here we developed an integrative approach to systematically identify and characterize non-coding regulatory variants with functional consequences in human hematopoietic malignancies. Combining targeted resequencing of hematopoietic lineage-specific CREs and mutation discovery, and uncovered 1,837 recurrently mutated CREs containing leukemia-associated non-coding variants. By enhanced CRISPR/dCas9-based CRE perturbation screening and functional analyses, we identified 218 variant-associated oncogenic or tumor suppressive CREs in human leukemia. Non-coding variants at KRAS and PER2 enhancers reside in nuclear receptor (NR) binding regions and modulate transcriptional activities in response to NR signaling in situ in leukemia cells. NR binding sites frequently co-localize with non-coding variants across cancer types. Hence, recurrent non-coding variants connect enhancer dysregulation with nuclear receptor signaling in hematopoietic malignancies. Overall design: ATAC-seq was performed to determine the chromatin accessibility in human leukemia cell lines

编码蛋白基因的突变已被广泛证实为人类癌症的发病基础，但占基因组相当比例且包含顺式调控元件（cis-regulatory elements, CREs）的非编码基因组区域发生改变，如何促进癌症病理生理进程，这一问题仍尚未明确。本研究开发了一套整合分析方法，以系统性鉴定并表征人类造血系统恶性肿瘤中具有功能效应的非编码调控变异。本研究结合造血谱系特异性顺式调控元件的靶向重测序与突变挖掘，共鉴定出1837个携带白血病相关非编码变异的复发性突变顺式调控元件。通过增强型CRISPR/dCas9介导的顺式调控元件扰动筛选与功能分析，本研究在人类白血病细胞中鉴定出218个与变异相关的致癌或抑癌顺式调控元件。KRAS与PER2增强子区域的非编码变异位于核受体（nuclear receptor, NR）结合区域内，可在白血病细胞中原位调控核受体信号通路应答相关的转录活性。在多种癌症类型中，核受体结合位点常与非编码变异共定位。因此，复发性非编码变异将造血系统恶性肿瘤中的增强子失调与核受体信号通路建立了关联。实验整体设计：本研究通过转座酶可及性测序分析（Assay for Transposase-Accessible Chromatin using sequencing, ATAC-seq）检测人类白血病细胞系的染色质可及性。