miR-135b, upregulated in breast cancer, promotes cell growth and disrupts the cell cycle by regulating LATS2
miR-135b, upregulated in breast cancer, promotes cell growth and disrupts the cell cycle by regulating LATS2
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miR-135b 在乳腺癌中表达上调，通过调节 LATS2 促进细胞生长并破坏细胞周期
DOI：
10.3892/ijo.2016.3405
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发表时间：
2016-05-01
影响因子：
5.2
通讯作者：
Fang, Lin
中科院分区：
文献类型：
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作者：
Hua, Kaiyao;Jin, Jiali;Fang, Lin
MicroRNAs调节失调(MiRNAs)在癌症进展中起着关键作用。在人类癌症中，它们既可以作为癌基因，也可以作为肿瘤抑制基因。本研究的目的是探讨miR-135B在乳腺癌中的重要作用，并验证miR-135B是否通过作用于HIPPO通路中的特定靶点来调节乳腺癌细胞的增殖。定量逆转录聚合酶链式反应(qRT-PCR)定量检测miR-135B在乳腺癌组织和细胞系中的表达水平。采用四甲基偶氮唑盐比色法、集落形成实验、细胞迁移实验、细胞侵袭实验和细胞周期实验对miR-135B的功能进行了研究。荧光素酶报告实验验证了miR-135B的一个可能的靶点的调控，并与蛋白质印迹分析进行了佐证。最后，我们验证了LATS2-siRNA转染后细胞功能的变化。我们的实验表明，miR-135B在乳腺癌标本和乳腺癌细胞中的表达普遍高于在癌旁正常组织和非恶性乳腺上皮细胞中的表达。MiR-135B的增强表达可以调节细胞的增殖、迁移和侵袭，并扰乱乳腺癌细胞的细胞周期。荧光素酶分析表明miR-135B直接与河马途径中的关键基因LATS2(大肿瘤抑制因子2)的3‘-非翻译区(3’-UTR)结合。Western印迹分析证实miR-135B在蛋白水平调控LATS2的表达。进一步研究表明，河马LATS2下游基因如细胞周期蛋白依赖性蛋白2(CDK2)和磷酸化相关蛋白(p-YAP)也可受miR-135B和LATS2轴的调控。下调内源性LATS2可以模拟miR-135B上调乳腺癌的结果。综上所述，我们的发现表明miR-135B和LATS2轴可能是未来乳腺癌的潜在治疗靶点。
Dysregulation of microRNAs (miRNAs) plays a critical role in cancer progression. They can act as either oncogenes or tumor suppressor genes in human cancer. The purpose of this study was to investigate the crucial role of miR-135b in breast cancer and to validate whether miR-135b could regulate proliferation of breast cancer cells by effecting specific targets in the Hippo pathway. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) was carried out to quantify the expression levels of miR-135b in both breast cancer tissues and cell lines. To characterize the function of miR-135b, MTT assays, colony formation assays, cell migration assays, cell invasion assays, and cell cycle assays were used. Luciferase reporter assays were performed to validate the regulation of a putative target of miR-135b, in corroboration with western blot assays. Finally, we verified the changes of cellular function after transfection of LATS2-siRNA. Our experiments indicate that expression of miR-135b was commonly upregulated in breast cancer specimens and breast cancer cells when compared with that in adjacent normal tissues and non-malignant breast epithelial cells. Enforced expression of miR-135b can regulate cellular proliferation, migration and invasion as well as disrupt the cell cycle of breast cancer cells. Luciferase assays revealed that miR-135b directly bound to the 3'-untranslated region (3'-UTR) of LATS2 (large tumor suppressor kinase 2), a critical gene in the Hippo pathway. Western blot analysis verified that miR-135b regulated the expression of LATS2 at protein levels. Further study demonstrated that the downstream gene of LATS2 in the Hippo pathway, such as cyclin-dependent kinase 2 (CDK2) and Phospho-Yes-associated protein (p-YAP), can also be regulated by miR-135b and LATS2 axis. Knockdown of endogenous LATS2 can mimic the result of miR-135b up-regulation in breast cancer. Taken together, our findings reveal that the miR-135b and LATS2 axis may be a potential therapeutic target for breast cancer in the future.