Functional Analysis of the 3' Untranslated Region of the Human Gene in Regulating Gene Expression in Vitro

Overview

Journal Neuropsychiatr Dis Treat

Publisher Dove Medical Press

Specialty Psychiatry

Date 2020 Oct 29

PMID 33116535

Citations 3

Authors

Yong-Ping Liu

Xue Wu

Jing-Hua Meng

Jun Yao

Bao-Jie Wang

Affiliations

Soon will be listed here.

Abstract

Purpose: Abnormal expression of the NR1 subunit of the N-methyl-d-aspartate (NMDA) receptor may potentially increase the susceptibility to neuropsychiatric diseases. The purpose of this study was to investigate the functional sequence of the 3'UTR of the human GRIN1 gene, which encodes the GluN1 receptor to determine the effect on the expression of GluN1 receptor.

Methods: We transferred seven recombinant pmirGLO recombinant vectors containing the 3'UTR truncated fragment of the GRIN1 gene into HEK-293, SK-N-SH, and U87 cell lines and compared the relative fluorescence intensity of adjacent length fragments. The TargetScan database was used to predict miRNAs. Then, miRNA mimics/inhibitors were co-transfected into the three cell lines with the 3'UTR of GRIN1 (pmirGLO - GRIN1), to investigate their influence on GRIN1 gene expression.

Results: Compared with the pmirGLo-Basic vector, the relative fluorescence intensity of the complete GRIN1 gene 3'UTR recombinant sequence -27 bp - +1284 bp (the next base of the stop codon is +1) was significantly decreased in all three cell lines. The relative fluorescence intensities were significantly different between -27 bp - +294 bp and -27 bp - +497 bp regions, and between -27 bp - +708 bp and -27 bp - +907 bp regions. According to the prediction of the TargetScan database and analysis, miR-212-5p, miR-324-3p and miR-326 may bind to +295 bp - +497 bp, while miR-491-5p may bind to +798 bp - +907 bp. After co-transfection of miRNA mimic/inhibitor or mimic/inhibitor NC with a recombinant vector in the 3'UTR region of GRIN1 gene, we found that has-miR-491-5p inhibited GRIN1 expression significantly in all three cell lines, while has-miR-326 inhibitor upregulated GRIN1 expression in HEK-293 and U87 cells.

Conclusion: miR-491-5p may bind to the 3'UTR of the GRIN1 gene (+799 bp - +805 bp, the next base of the stop codon is +1) and down-regulate gene expression in HEK-293, SK-N-SH, and U87 cell lines, which implicates a potential role of miR-491-5p in central nervous system diseases.

Citing Articles

A Scoping Review of Rodent Studies Investigating the Epigenetic Mechanisms in the Brain Underlying the Effects of Diet on Depressive-like Behaviour.

Sanchez-Lafuente C, Reive B, Kalynchuk L, Caruncho H Biomedicines. 2022; 10(12).

PMID: 36551970 PMC: 9775686. DOI: 10.3390/biomedicines10123213.

Maternal High-Fat Diet Modulates Gene Expression in Male Rat Offspring.

Gawlinski D, Gawlinska K, Smaga I Nutrients. 2021; 13(8).

PMID: 34445045 PMC: 8402185. DOI: 10.3390/nu13082885.

Dopamine System, NMDA Receptor and EGF Family Expressions in Brain Structures of Bl6 and 129Sv Strains Displaying Different Behavioral Adaptation.

Varul J, Eskla K, Piirsalu M, Innos J, Philips M, Visnapuu T Brain Sci. 2021; 11(6).

PMID: 34072341 PMC: 8227283. DOI: 10.3390/brainsci11060725.

References

Kristiansen L, Huerta I, Beneyto M, Meador-Woodruff J . NMDA receptors and schizophrenia. Curr Opin Pharmacol. 2006; 7(1):48-55. DOI: 10.1016/j.coph.2006.08.013. View

Kalev-Zylinska M, Symes W, Young D, During M . Knockdown and overexpression of NR1 modulates NMDA receptor function. Mol Cell Neurosci. 2009; 41(4):383-96. PMC: 3727896. DOI: 10.1016/j.mcn.2009.04.003. View

Lu L, Cai M, Peng M, Wang F, Zhai X . miR-491-5p functions as a tumor suppressor by targeting IGF2 in colorectal cancer. Cancer Manag Res. 2019; 11:1805-1816. PMC: 6391127. DOI: 10.2147/CMAR.S183085. View

Gonzalez-Arenas A, De La Fuente-Granada M, Camacho-Arroyo I, Zamora-Sanchez C, Pina-Medina A, Segura-Uribe J . Tibolone Effects on Human Glioblastoma Cell Lines. Arch Med Res. 2019; 50(4):187-196. DOI: 10.1016/j.arcmed.2019.08.001. View

He B, Chen W, Zeng J, Tong W, Zheng P . MicroRNA-326 decreases tau phosphorylation and neuron apoptosis through inhibition of the JNK signaling pathway by targeting VAV1 in Alzheimer's disease. J Cell Physiol. 2019; 235(1):480-493. DOI: 10.1002/jcp.28988. View

Biedler J, Helson L, Spengler B . Morphology and growth, tumorigenicity, and cytogenetics of human neuroblastoma cells in continuous culture. Cancer Res. 1973; 33(11):2643-52. View

Akhtar M, Micolucci L, Islam M, Olivieri F, Procopio A . A Practical Guide to miRNA Target Prediction. Methods Mol Biol. 2019; 1970:1-13. DOI: 10.1007/978-1-4939-9207-2_1. View

Ibrahim H, Hogg Jr A, Healy D, Haroutunian V, Davis K, Meador-Woodruff J . Ionotropic glutamate receptor binding and subunit mRNA expression in thalamic nuclei in schizophrenia. Am J Psychiatry. 2000; 157(11):1811-23. DOI: 10.1176/appi.ajp.157.11.1811. View

Dos Santos M, Barreto-Sanz M, Correia B, Bell R, Widnall C, Perez L . miRNA-based signatures in cerebrospinal fluid as potential diagnostic tools for early stage Parkinson's disease. Oncotarget. 2018; 9(25):17455-17465. PMC: 5915128. DOI: 10.18632/oncotarget.24736. View

10.

Liu R, Li F, Zhao W . Long noncoding RNA NEAT1 knockdown inhibits MPP-induced apoptosis, inﬂammation and cytotoxicity in SK-N-SH cells by regulating miR-212-5p/RAB3IP axis. Neurosci Lett. 2020; 731:135060. DOI: 10.1016/j.neulet.2020.135060. View

11.

Shukla G, Singh J, Barik S . MicroRNAs: Processing, Maturation, Target Recognition and Regulatory Functions. Mol Cell Pharmacol. 2012; 3(3):83-92. PMC: 3315687. View

12.

Wu X, Ding M, Liu Y, Xia X, Xu F, Yao J . hsa-miR-3177-5p and hsa-miR-3178 Inhibit 5-HT1A Expression by Binding the 3'-UTR Region . Front Mol Neurosci. 2019; 12:13. PMC: 6365703. DOI: 10.3389/fnmol.2019.00013. View

13.

Herrera-Espejo S, Santos-Zorrozua B, Alvarez-Gonzalez P, Lopez-Lopez E, Garcia-Orad A . A Systematic Review of MicroRNA Expression as Biomarker of Late-Onset Alzheimer's Disease. Mol Neurobiol. 2019; 56(12):8376-8391. DOI: 10.1007/s12035-019-01676-9. View

14.

Yu T, Wang L, Li W, Zuo Q, Li M, Zou Q . Downregulation of miR-491-5p promotes gastric cancer metastasis by regulating SNAIL and FGFR4. Cancer Sci. 2018; 109(5):1393-1403. PMC: 5980274. DOI: 10.1111/cas.13583. View

15.

Singh A, Sen D . MicroRNAs in Parkinson's disease. Exp Brain Res. 2017; 235(8):2359-2374. DOI: 10.1007/s00221-017-4989-1. View

16.

Annese A, Manzari C, Lionetti C, Picardi E, Horner D, Chiara M . Whole transcriptome profiling of Late-Onset Alzheimer's Disease patients provides insights into the molecular changes involved in the disease. Sci Rep. 2018; 8(1):4282. PMC: 5844946. DOI: 10.1038/s41598-018-22701-2. View

17.

Chen D, Li J, Li S, Han P, Li N, Wang Y . miR-184 promotes cell proliferation in tongue squamous cell carcinoma by targeting SOX7. Oncol Lett. 2018; 16(2):2221-2228. PMC: 6036414. DOI: 10.3892/ol.2018.8906. View

18.

Xu Y, Hou R, Lu Q, Zhang Y, Chen L, Zheng Y . MiR-491-5p negatively regulates cell proliferation and motility by targeting PDGFRA in prostate cancer. Am J Cancer Res. 2018; 7(12):2545-2553. PMC: 5752694. View

19.

Rupaimoole R, Slack F . MicroRNA therapeutics: towards a new era for the management of cancer and other diseases. Nat Rev Drug Discov. 2017; 16(3):203-222. DOI: 10.1038/nrd.2016.246. View

20.

Papouin T, Oliet S . Organization, control and function of extrasynaptic NMDA receptors. Philos Trans R Soc Lond B Biol Sci. 2014; 369(1654):20130601. PMC: 4173287. DOI: 10.1098/rstb.2013.0601. View