Bioengineered miR-34a modulates mitochondrial inner membrane protein 17 like 2 (MPV17L2) expression toward the control of cancer cell mitochondrial functions.
Bioengineered miR-34a modulates mitochondrial inner membrane protein 17 like 2 (MPV17L2) expression toward the control of cancer cell mitochondrial functions.
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DOI：
10.1080/21655979.2022.2076399
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发表时间：
2022-05
期刊：
影响因子：
4.9
通讯作者：
中科院分区：
文献类型：
--
作者：
基因组衍生的microRNAs （miRNAs或miRs）控制转录后基因表达，对各种细胞过程至关重要。最近，我们发明了一种新的平台技术，可以实现高产量的全人源化生物工程miRNA制剂（hBERAs）的研究和开发。本研究旨在生产和利用一种新的生物miR-34a-5p（或miR-34a）分子，即hBERA/miR-34a，来描述miR-34a-5p在人类癌细胞线粒体功能调控中的作用。生物工程hBERA/miR-34a通过体内发酵生产，通过阴离子交换快速蛋白液相色谱纯化。通过选择性茎环反转录定量聚合酶链式反应分析确定hEBRA/miR-34a在人骨肉瘤和肺癌细胞中靶向miR-34a-5p。通过双荧光素酶报告基因检测，证实线粒体内膜蛋白MPV17样2 （MPV17L2）是miR-34a-5p的直接靶标。Western blot分析显示，生物工程miR-34a-5p有效降低了MPV17L2蛋白的结果，导致呼吸链复合体I活性和细胞内ATP水平大大降低，这是用特定的检测试剂盒测定的。此外，我们进行了海马Mito应激试验，结果显示生物miR-34a-5p显著降低了癌细胞线粒体呼吸能力，同时氧化应激显著增加，凋亡细胞死亡率升高，分别表现为活性氧和选择性凋亡生物标志物水平升高。这些结果证明了miR-34a-5p-MPV17L2通路在人类癌细胞线粒体功能控制中的存在和参与，并支持了新型生物工程miRNA分子在功能研究中的应用。
Genome-derived microRNAs (miRNAs or miRs) control post-transcriptional gene expression critical for various cellular processes. Recently, we have invented a novel platform technology to achieve high-yield production of fully humanized, bioengineered miRNA agents (hBERAs) for research and development. This study is aimed to produce and utilize a new biologic miR-34a-5p (or miR-34a) molecule, namely, hBERA/miR-34a, to delineate the role of miR-34a-5p in the regulation of mitochondrial functions in human carcinoma cells. Bioengineered hBERA/miR-34a was produced through in vivo fermentation production and purified by anion exchange fast protein liquid chromatography. hEBRA/miR-34a was processed to target miR-34a-5p in human osteosarcoma and lung cancer cells, as determined by selective stem-loop reverse transcription quantitative polymerase chain reaction analysis. The mitochondrial inner membrane protein MPV17 like 2 (MPV17L2) was validated as a direct target for miR-34a-5p by dual luciferase reporter assay. Western blot analysis revealed that bioengineered miR-34a-5p effectively reduced MPV17L2 protein outcomes, leading to much lower levels of respiratory chain Complex I activities and intracellular ATP that were determined with specific assay kits. Moreover, Seahorse Mito Stress Test assay was conducted, and the results showed that biologic miR-34a-5p sharply reduced cancer cell mitochondrial respiration capacity, accompanied by a remarkable increase of oxidative stress and elevated apoptotic cell death, which are manifested by greater levels of reactive oxygen species and selective apoptosis biomarkers, respectively. These results demonstrate the presence and involvement of the miR-34a-5p-MPV17L2 pathway in the control of mitochondrial functions in human carcinoma cells and support the utility of novel bioengineered miRNA molecules for functional studies.
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影响因子：
5.5
作者：
Geldon S;Fernández-Vizarra E;Tokatlidis K
通讯作者：
Tokatlidis K
DOI：
10.1016/j.apsb.2021.07.027
发表时间：
2021-12
期刊：
Acta pharmaceutica Sinica. B
影响因子：
--
作者：
Deng L;Petrek H;Tu MJ;Batra N;Yu AX;Yu AM
通讯作者：
Yu AM
影响因子：
4.8
作者：
Antonenkov, Vasily D.;Isomursu, Antti;Hiltunen, J. Kalervo
通讯作者：
Hiltunen, J. Kalervo
影响因子：
29
作者：
Guerrero-Castillo, Sergio;Baertling, Fabian;Nijtmans, Leo
通讯作者：
Nijtmans, Leo
影响因子：
10.3
作者：
Cortez, Maria Angelica;Ivan, Cristina;Welsh, James W.
通讯作者：
Welsh, James W.