Oncogenic lncRNAs alter epigenetic memory at a fragile chromosomal site in human cancer cells [WGS]

Chromosome instability is a key event in cancer progression. The essential histone H3 variant CENP-A plays a fundamental role in defining centromere identity, structure, and function, but is innately overexpressed in several types of solid cancers. In the cancer background, excess CENP-A is deposited ectopically on chromosome arms, including at the 8q24/cMYC locus, by invading transcription-coupled H3.3 chaperone pathways. Intriguingly, in many cancers, transcription of lncRNAs is upregulated and correlates with poor prognosis, therapeutic resistance, and cancer recurrence in patients. Here, we report that the transcription of chromosome 8q24-derived oncogenic lncRNAs plays an unanticipated role in altering the chromatin landscape of the 8q24 locus. We report that transcription of oncogenic ncRNAs and associated R-loop formation at the 8q24 genomic locus results in the recruitment of H3.3 chaperone-CENP-A histone variant complexes to 8q24. Finally, we demonstrate that a transgene cassette which encodes a specific oncogenic lncRNA from the 8q24 region integrated into a naïve chromosome locus, recruits CENP-A to the new location specifically in a cis-acting manner. These data provide a plausible mechanistic link between locus-specific oncogenic lncRNAs, aberrant local chromatin structure, and the generation of new epigenetic memory in human cancer cells. Whole Genome Sequencing of human SW480 colon cancer cells knocked-in with PCAT2 gene array (3x) under CMV promoter in replicates. gDNA was isolated with PCI method. The samples quality was assessed by BioAnalyzer.

染色体不稳定（chromosome instability）是癌症进展中的关键事件。核心组蛋白H3变体着丝粒蛋白A（CENP-A）在定义着丝粒的身份、结构与功能方面发挥着基础性作用，但其在多种实体瘤中本就存在过表达现象。在癌症背景下，过量的CENP-A会通过入侵转录偶联的H3.3分子伴侣通路，异位沉积在染色体臂上，包括8q24/cMYC基因座。
有趣的是，在多种癌症中，长链非编码RNA（long non-coding RNAs, lncRNAs）的转录水平会上调，且与患者预后不良、治疗耐药及癌症复发密切相关。本研究发现，染色体8q24区域来源的致癌性lncRNAs的转录，在重塑8q24基因座的染色质景观方面发挥了此前未被认知的作用。本研究还证实，致癌性非编码RNA的转录以及8q24基因组位点处伴随形成的R环（R-loop）结构，会将H3.3分子伴侣-着丝粒蛋白A（CENP-A）组蛋白变体复合物招募至8q24区域。最后，本研究证明，将一段编码8q24区域特定致癌性lncRNA的转基因盒（transgene cassette）整合至天然染色体位点后，能够以顺式作用（cis-acting）的方式特异性地将CENP-A招募至新位点。
上述数据为位点特异性致癌性lncRNAs、异常的局部染色质结构，与人类癌细胞中新表观遗传记忆（epigenetic memory）的产生之间，搭建了一条合理的机制性关联。
本数据集来自于在巨细胞病毒（Cytomegalovirus, CMV）启动子下敲入3倍PCAT2基因阵列的人类SW480结肠癌细胞的全基因组测序，设置生物学重复。采用PCI法分离基因组DNA（genomic DNA, gDNA），并通过生物分析仪（BioAnalyzer）评估样本质量。