Nie Yuanyuan, Zhou Lei, Wang Hong, Chen Naifei, Jia Lin, Wang Cong, Wang Yichen, Chen Jingcheng, Wen Xue, Niu Chao, Li Hui, Guo Rui, Zhang Songling, Cui Jiuwei, Hoffman Andrew R, Hu Ji-Fan, Li Wei
Stem Cell and Cancer Center, First Hospital, Jilin University Changchun 130061, Jilin, China.
Stanford University Medical School, VA Palo Alto Health Care System Palo Alto, CA 94304, USA.
Am J Cancer Res. 2019 Aug 1;9(8):1635-1649. eCollection 2019.
RUNX1 is frequently mutated as chromosomal translocations in a variety of hematological malignancies. Recent studies show that RUNX1 is also mutated somatically in many solid tumors. We have recently identified a 260 kb un-spliced intragenic overlapping long noncoding RNA RUNXOR in the RUNX1 locus, yet its role as an epigenetic regulator in tumors remains to be characterized. To delineate this RUNXOR-RUNX1 regulatory interplay in breast cancer cells, we devised a novel "gene in situ cis-activation" approach to activate the endogenous RUNXOR gene. We found that the in situ activation of RUNXOR lncRNA upregulated RUNX1 in cis from the P1 promoter. The preferred activation of the P1 promoter caused a shift to the RUNX1c isoform expression. Using a chromatin conformation capture (3C) approach, we showed that RUNXOR lncRNA epigenetically activated the RUNX1 P1 promoter in cis by altering the local chromatin structure. The binding of RUNXOR lncRNA triggered DNA demethylation and induced active histone modification markers in the P1 CpG island. Changes in RUNX1 isoform composition correlated with a trend to cell cycle arrest at G0/G1, although cell proliferation rate, apoptosis, and migration ability were not significantly changed. Our results reveal an underlying epigenetic mechanism by which the lncRNA regulates in cis the RUNX1 promoter usage in breast cancer cells, thereby shedding light on potential genetic therapies in malignancies in which RUNX1 loss-of-function mutations frequently occur.
RUNX1在多种血液系统恶性肿瘤中常因染色体易位而发生突变。最近的研究表明，RUNX1在许多实体瘤中也存在体细胞突变。我们最近在RUNX1基因座中鉴定出一个260 kb未剪接的基因内重叠长链非编码RNA RUNXOR，但其在肿瘤中作为表观遗传调节因子的作用仍有待确定。为了阐明乳腺癌细胞中这种RUNXOR-RUNX1的调控相互作用，我们设计了一种新颖的“基因原位顺式激活”方法来激活内源性RUNXOR基因。我们发现RUNXOR lncRNA的原位激活从P1启动子顺式上调了RUNX1。P1启动子的优先激活导致了向RUNX1c异构体表达的转变。使用染色质构象捕获（3C）方法，我们表明RUNXOR lncRNA通过改变局部染色质结构在顺式上表观遗传激活了RUNX1 P1启动子。RUNXOR lncRNA的结合触发了DNA去甲基化，并在P1 CpG岛中诱导了活性组蛋白修饰标记。RUNX1异构体组成的变化与细胞周期在G0/G1期停滞的趋势相关，尽管细胞增殖率、凋亡和迁移能力没有明显变化。我们的结果揭示了一种潜在的表观遗传机制，lncRNA通过该机制在顺式上调节乳腺癌细胞中RUNX1启动子的使用，从而为经常发生RUNX1功能丧失突变的恶性肿瘤的潜在基因治疗提供了线索。
