Fundación Ciencia & Vida/Andes Biotechnologies SpA, 7780272, Santiago, Chile.
Department of Biological Sciences, Faculty of Life Sciences, Universidad Andrés Bello, 8370134, Santiago, Chile.
Cell Death Dis. 2019 May 29;10(6):423. doi: 10.1038/s41419-019-1649-3.
The family of long noncoding mitochondrial RNAs (ncmtRNAs), comprising sense (SncmtRNA), and antisense (ASncmtRNA-1 and ASncmtRNA-2) members, are differentially expressed according to cell proliferative status; SncmtRNA is expressed in all proliferating cells, while ASncmtRNAs are expressed in normal proliferating cells, but is downregulated in tumor cells. ASncmtRNA knockdown with an antisense oligonucleotide induces massive apoptosis in tumor cell lines, without affecting healthy cells. Apoptotic death is preceded by proliferation blockage, suggesting that these transcripts are involved in cell cycle regulation. Here, we show that ASncmtRNA knockdown induces cell death preceded by proliferative blockage in three different human breast cancer cell lines. This effect is mediated by downregulation of the key cell cycle progression factors cyclin B1, cyclin D1, CDK1, CDK4, and survivin, the latter also constituting an essential inhibitor of apoptosis, underlying additionally the onset of apoptosis. The treatment also induces an increase in the microRNA hsa-miR-4485-3p, whose sequence maps to ASncmtRNA-2 and transfection of MDA-MB-231 cells with a mimic of this miRNA induces cyclin B1 and D1 downregulation. Other miRNAs that are upregulated include nuclear-encoded hsa-miR-5096 and hsa-miR-3609, whose mimics downregulate CDK1. Our results suggest that ASncmtRNA targeting blocks tumor cell proliferation through reduction of essential cell cycle proteins, mediated by mitochondrial and nuclear miRNAs. This work adds to the elucidation of the molecular mechanisms behind cell cycle arrest preceding tumor cell apoptosis induced by ASncmtRNA knockdown. As proof-of-concept, we show that in vivo knockdown of ASncmtRNAs results in drastic inhibition of tumor growth in a xenograft model of MDA-MB-231 subcutaneous tumors, further supporting this approach for the development of new therapeutic strategies against breast cancer.
长非编码线粒体 RNA（ncmtRNA）家族，包括 sense（SncmtRNA）和 antisense（ASncmtRNA-1 和 ASncmtRNA-2）成员，根据细胞增殖状态呈现不同的表达模式；SncmtRNA 在所有增殖细胞中表达，而 ASncmtRNAs 在正常增殖细胞中表达，但在肿瘤细胞中下调。用反义寡核苷酸敲低 ASncmtRNA 会诱导肿瘤细胞系发生大量凋亡，而不影响健康细胞。凋亡死亡之前有增殖阻断，表明这些转录物参与细胞周期调控。在这里，我们显示 ASncmtRNA 敲低会在三种不同的人乳腺癌细胞系中诱导增殖阻断之前的细胞死亡。这种效应是通过下调关键细胞周期进程因子 cyclin B1、cyclin D1、CDK1、CDK4 和 survivin 介导的，后者也是凋亡的必需抑制剂，此外还引发了凋亡的发生。该治疗还会引起 microRNA hsa-miR-4485-3p 的增加，其序列与 ASncmtRNA-2 映射，并且用该 miRNA 的模拟物转染 MDA-MB-231 细胞会导致 cyclin B1 和 D1 的下调。其他上调的 miRNA 包括核编码的 hsa-miR-5096 和 hsa-miR-3609，其模拟物下调 CDK1。我们的结果表明，ASncmtRNA 靶向通过减少必需的细胞周期蛋白，通过线粒体和核 miRNA 介导，阻断肿瘤细胞增殖。这项工作增加了对 ASncmtRNA 敲低诱导肿瘤细胞凋亡之前细胞周期阻滞的分子机制的阐明。作为概念验证，我们表明在 MDA-MB-231 皮下肿瘤的异种移植模型中，ASncmtRNAs 的体内敲低导致肿瘤生长的急剧抑制，进一步支持了这种方法用于开发针对乳腺癌的新治疗策略。
