The highly expressed 5'isomiR of hsa-miR-140-3p contributes to the tumor-suppressive effects of miR-140 by reducing breast cancer proliferation and migration.
The highly expressed 5'isomiR of hsa-miR-140-3p contributes to the tumor-suppressive effects of miR-140 by reducing breast cancer proliferation and migration.
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DOI：
10.1186/s12864-016-2869-x
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发表时间：
2016-08-08
期刊：
影响因子：
4.4
通讯作者：
Körner C
中科院分区：
文献类型：
--
作者：
Salem O;Erdem N;Jung J;Münstermann E;Wörner A;Wilhelm H;Wiemann S;Körner C
mirna是一种小的非编码RNA分子，在基因表达的转录后调控中起重要作用。相同miRNA的长度和/或序列变体称为同分异构体。虽然大多数同分异构体与它们的标准对应物相比在功能上是冗余的，但所谓的5 ‘同分异构体表现出移位的5 ’端，因此移位的种子序列导致不同的靶谱。然而，对这些同工异构体的功能相关性知之甚少。对乳腺癌细胞系的miRNA-seq数据进行分析，鉴定出6对高表达的mirna和相关的5 ' isomir。其中，hsa-miR-140-3p特别令人感兴趣，因为它的5'isomiR比miRbase中注释的标准miRNA表达更高。该miRNA先前已被证明可以控制乳腺癌细胞的干细胞性。乳腺癌患者的miRNAseq数据（TCGA数据集）显示，与正常乳腺组织相比，标准hsa-miR-140-3p及其5'isomiR-140-3p在患者肿瘤中均高表达。在目前的工作中，我们提出了5'isomiR-140-3p的功能表征及其在MCF10A、MDA-MB-468和MDA-MB-231细胞系中与标准hsa-miR-140-3p过表达相关的细胞表型。与标准hsa-miR-140-3p的作用相反，过表达5'isomiR-140-3p导致细胞活力下降。后一种观察结果得到了细胞周期分析的支持，其中5'isomiR-140-3p而不是hsa-miR-140-3p导致细胞周期阻滞在G0/ g1期。此外，发现5'ismoiR-140-3p过表达导致三种细胞系的细胞迁移减少。我们发现了3个新的5'isomiR-140-3p的直接靶基因；COL4A1， ITGA6和marksl1。最后，我们已经证明，敲低这些基因部分地表型化了5'isomiR-140-4p过表达的影响，其中COL4A1和ITGA6敲低导致细胞活力降低和细胞周期停滞，而MARCKSL1敲低导致细胞迁移潜力降低。总之，这项工作提供了证据，证明在标准的hsa-miR-140-3p和新发现的5'isomiR-140-3p之间存在功能协同作用，通过同时靶向与分化、增殖和迁移相关的基因来抑制乳腺癌的生长和进展。本文的在线版本（doi:10.1186/s12864-016-2869-x）包含补充材料，授权用户可以使用。
miRNAs are small noncoding RNA molecules that play an important role in post-transcriptional regulation of gene expression. Length and/or sequence variants of the same miRNA are termed isomiRs. While most isomiRs are functionally redundant compared to their canonical counterparts, the so-called 5’isomiRs exhibit a shifted 5’ end and therefore a shifted seed sequence resulting in a different target spectrum. However, not much is known about the functional relevance of these isoforms. Analysis of miRNA-seq data from breast cancer cell lines identified six pairs of highly expressed miRNAs and associated 5’isomiRs. Among them, hsa-miR-140-3p was of particular interest because its 5’isomiR showed higher expression compared to the canonical miRNA annotated in miRbase. This miRNA has previously been shown to control stemness of breast cancer cells. miRNAseq data of breast cancer patients (TCGA dataset) showed that both the canonical hsa-miR-140-3p and its 5’isomiR-140-3p were highly expressed in patients’ tumors compared to normal breast tissue. In the current work, we present the functional characterization of 5’isomiR-140-3p and the cellular phenotypes associated with its overexpression in MCF10A, MDA-MB-468 and MDA-MB-231 cell lines in comparison to the canonical hsa-miR-140-3p. Contrary to the effect of the canonical hsa-miR-140-3p, overexpression of the 5’isomiR-140-3p led to a decrease in cell viability. The latter observation was supported by cell cycle analysis, where the 5’isomiR-140-3p but not the hsa-miR-140-3p caused cell cycle arrest in G0/G1-phase. Additionally, 5’ismoiR-140-3p overexpression was found to cause a decrease in cell migration in the three cell lines. We identified three novel direct target genes of the 5’isomiR-140-3p; COL4A1, ITGA6 and MARCKSL1. Finally, we have shown that knocking down these genes partially phenocopied the effects of the 5’isomiR-140-4p overexpression, where COL4A1 and ITGA6 knockdown led to reduced cell viability and cell cycle arrest, while MARCKSL1 knockdown resulted in a decrease in the migratory potential of cells. In summary, this work presents evidence that there is functional synergy between the canonical hsa-miR-140-3p and the newly identified 5’isomiR-140-3p in suppressing growth and progression of breast cancer by simultaneously targeting genes related to differentiation, proliferation, and migration. The online version of this article (doi:10.1186/s12864-016-2869-x) contains supplementary material, which is available to authorized users.