RNA-Binding Protein MSI2 Binds to miR-301a-3p and Facilitates Its Distribution in Mitochondria of Endothelial Cells

Qian Qian Guo; Jing Gao; Xiao Wei Wang; Xian Lun Yin; Shu Cui Zhang; Xue Li; Lian Li Chi; Xiao Ming Zhou; Zhe Wang; Qun Ye Zhang

doi:10.3389/fmolb.2020.609828

RNA-Binding Protein MSI2 Binds to miR-301a-3p and Facilitates Its Distribution in Mitochondria of Endothelial Cells

Front Mol Biosci. 2021 Jan 21:7:609828. doi: 10.3389/fmolb.2020.609828. eCollection 2020.

Authors

Qian Qian Guo^{1

2

3

4}, Jing Gao^{1

2

3}, Xiao Wei Wang^{1

2

3}, Xian Lun Yin^{1

2

3}, Shu Cui Zhang^{1

2

3}, Xue Li⁵, Lian Li Chi⁵, Xiao Ming Zhou^{6

7}, Zhe Wang^{8

9}, Qun Ye Zhang^{1

2

3

4}

Affiliations

¹ The Key Laboratory of Cardiovascular Remodeling and Function Research, Chinese Ministry of Education and Chinese National Health Commission, Shandong University, Jinan, China.
² Department of Cardiology, Cheeloo College of Medicine, Qilu Hospital, Shandong University, Jinan, China.
³ The State and Shandong Province Joint Key Laboratory of Translational Cardiovascular Medicine, Jinan, China.
⁴ Cardiovascular Disease Research Center of Shandong First Medical University, Central Hospital Affiliated to Shandong First Medical University, Jinan, China.
⁵ National Glycoengineering Research Center, Shandong University, Qingdao, China.
⁶ Institute of Endocrinology, Shandong Academy of Clinical Medicine, Jinan, China.
⁷ Shandong Clinical Medical Center of Endocrinology and Metabolism, Jinan, China.
⁸ Division of Endocrinology and Metabolism, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, China.
⁹ Division of Geriatrics, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, China.

Abstract

Numerous miRNAs have been detected in mitochondria, which play important roles in many physiological and pathophysiological processes. However, the dynamic changes of miRNA distribution in mitochondria and their mechanisms in reactive oxygen species (ROS)-induced endothelial injury remain unclear. Therefore, miRNA levels in whole cells and mitochondria of H₂O₂-treated endothelial cells were analyzed by small RNA sequencing in the present study. The results showed that H₂O₂ significantly reduced the relative mitochondrial distribution of dozens of miRNAs in human umbilical vein endothelial cells (HUVECs). Among the high-abundance miRNAs, miR-301a-3p has the most significant changes in the redistribution between cytosol and mitochondria confirmed by absolute quantitative polymerase chain reaction (qPCR). To unravel the mechanism of miR-301a-3p distribution in mitochondria, RNA pull-down followed by label-free quantitative proteomic analysis was performed, and RNA-binding protein Musashi RNA binding protein 2 (MSI2) was found to specifically bind to miR-301a-3p. Western blotting and immunofluorescence colocalization assay showed that MSI2 was located in mitochondria of various cell types. H₂O₂ significantly downregulated MSI2 expression in whole endothelial cells, promoted the distribution of MSI2 in cytosol and decreased its distribution in the mitochondria. Moreover, overexpression of MSI2 increased the mitochondrial distribution of miR-301a-3p, whereas inhibition of MSI2 decreased its distribution in mitochondria. Thus, MSI2 might be responsible for the distribution of miR-301a-3p between cytosol and mitochondria in endothelial cells. Our findings revealed for the first time that MSI2 was involved in the regulation of miRNA distribution in mitochondria and provided valuable insight into the mechanism of mitochondrial distribution of miRNAs.

Keywords: Musashi RNA binding protein 2; RNA-binding protein; endothelial cell injury; mitochondrial miRNA distribution; reactive oxygen species.