Login Register Cart 0
Research Article

P68 RNA Helicase (DDX5) Required for the Formation of Various Specific and Mature miRNA Active RISC Complexes

Author(s): Mariette Kokolo and Montse Bach-Elias*

Volume 11, Issue 1, 2022

Published on: 05 April, 2022

Page: [36 - 44] Pages: 9

DOI: 10.2174/2211536611666220218121640

open access plus

conference banner
Abstract

Introduction: DEAD-box RNA helicases catalyze the ATP-dependent unwinding of doublestranded RNA. In addition, they are required for protein displacement and remodelling of RNA or RNA/protein complexes. P68 RNA helicase regulates the alternative splicing of the important protooncogene H-Ras, and numerous studies have shown that p68 RNA helicase is probably involved in miRNA biogenesis, mainly through Drosha and RISC/DICER complexes.

Objective: This study aimed to determine how p68 RNA helicase affects the activity of selected mature miRNAs, including miR-342, miR-330, miR-138 and miR-206, miR-126, and miR-335, and let-7a, which are known to be related to cancer processes.

Methods: The miRNA levels were analyzed in stable HeLa cells containing p68 RNA helicase RNAi induced by doxycycline (DOX). Relevant results were repeated using transient transfection with pSuper/ pSuper-p68 RNA helicase RNAi to avoid DOX interference.

Results: Herein, we reported that p68 RNA helicase downregulation increases the accumulation of the mature miRNAs, such as miR-126, let-7a, miR-206, and miR-138. Interestingly, the accumulation of these mature miRNAs does not downregulate their known protein targets, thus suggesting that p68 RNA helicase is required for mature miRNA-active RISC complex activity.

Conclusion: Furthermore, we demonstrated that this requirement is conserved, as drosophila p68 RNA helicase can complete the p68 RNA helicase depleted activity in human cells. Dicer and Drosha proteins are not affected by the downregulation of p68 RNA helicase despite the fact that Dicer is also localized in the nucleus when p68 RNA helicase activity is reduced.

Keywords: p68 RNA helicase, miRNAs, mature miRNA active RISC complexes, miR-206, Dicer, Let-7a.

« Previous Next »
Graphical Abstract

[1]
Bleichert F, Baserga SJ. The long unwinding road of RNA helicases. Mol Cell 2007; 27(3): 339-52.
[http://dx.doi.org/10.1016/j.molcel.2007.07.014] [PMID: 17679086]
[2]
Sergeeva O, Zatsepin T. RNA helicases as shadow modulators of cell cycle progression. Int J Mol Sci 2021; 22(6): 2984.
[http://dx.doi.org/10.3390/ijms22062984] [PMID: 33804185]
[3]
Iggo RD, Lane DP. Nuclear protein p68 is an RNA-dependent ATPase. EMBO J 1989; 8(6): 1827-31.
[http://dx.doi.org/10.1002/j.1460-2075.1989.tb03577.x] [PMID: 2527746]
[4]
Cheng W, Chen G, Jia H, He X, Jing Z. DDX5 RNA helicases: Emerging roles in viral infection. Int J Mol Sci 2018; 19(4): 1122.
[http://dx.doi.org/10.3390/ijms19041122] [PMID: 29642538]
[5]
Fuller-Pace FV. The DEAD box proteins DDX5 (p68) and DDX17 (p72): multi-tasking transcriptional regulators. Biochim Biophys Acta 2013; 1829(8): 756-63.
[http://dx.doi.org/10.1016/j.bbagrm.2013.03.004] [PMID: 23523990]
[6]
Clark EL, Coulson A, Dalgliesh C, et al. The RNA helicase p68 is a novel androgen receptor coactivator involved in splicing and is overex-pressed in prostate cancer. Cancer Res 2008; 68(19): 7938-46.
[http://dx.doi.org/10.1158/0008-5472.CAN-08-0932] [PMID: 18829551]
[7]
Shin S, Rossow KL, Grande JP, Janknecht R. Involvement of RNA helicases p68 and p72 in colon cancer. Cancer Res 2007; 67(16): 7572-8.
[http://dx.doi.org/10.1158/0008-5472.CAN-06-4652] [PMID: 17699760]
[8]
Causevic M, Hislop RG, Kernohan NM, et al. Overexpression and poly-ubiquitylation of the DEAD-box RNA helicase p68 in colorectal tumours. Oncogene 2001; 20(53): 7734-43.
[http://dx.doi.org/10.1038/sj.onc.1204976] [PMID: 11753651]
[9]
Mooney SM, Grande JP, Salisbury JL, Janknecht R. Sumoylation of p68 and p72 RNA helicases affects protein stability and transactivation potential. Biochemistry 2010; 49(1): 1-10.
[http://dx.doi.org/10.1021/bi901263m] [PMID: 19995069]
[10]
Ali MAM. The DEAD-box protein family of RNA helicases: sentinels for a myriad of cellular functions with emerging roles in tumorigene-sis. Int J Clin Oncol 2021; 26(5): 795-825.
[http://dx.doi.org/10.1007/s10147-021-01892-1] [PMID: 33656655]
[11]
Guil S, de La Iglesia N, Fernández-Larrea J, et al. Alternative splicing of the human proto-oncogene c-H-ras renders a new Ras family pro-tein that trafficks to cytoplasm and nucleus. Cancer Res 2003; 63(17): 5178-87.
[PMID: 14500341]
[12]
Guil S, Gattoni R, Carrascal M, Abián J, Stévenin J, Bach-Elias M. Roles of hnRNP A1, SR proteins, and p68 helicase in c-H-ras alternative splicing regulation. Mol Cell Biol 2003; 23(8): 2927-41.
[http://dx.doi.org/10.1128/MCB.23.8.2927-2941.2003] [PMID: 12665590]
[13]
Camats M, Guil S, Kokolo M, Bach-Elias M. P68 RNA helicase (DDX5) alters activity of cis- and trans-acting factors of the alternative splicing of H-Ras. PLoS One 2008; 3(8): e2926.
[http://dx.doi.org/10.1371/journal.pone.0002926] [PMID: 18698352]
[14]
Camats M, Kokolo M, Heesom KJ, Ladomery M, Bach-Elias M. P19 H-ras induces G1/S phase delay maintaining cells in a reversible qui-escence state. PLoS One 2009; 4(12): e8513.
[http://dx.doi.org/10.1371/journal.pone.0008513] [PMID: 20046837]
[15]
Kokolo M, Bach-Elias M. Downregulation of p68 RNA helicase (DDX5) activates a survival pathway involving mTOR and MDM2 signals. Folia Biol (Praha) 2017; 63(2): 52-9.
[PMID: 28557706]
[16]
Bartel DP. MicroRNAs: genomics, biogenesis, mechanism, and function. Cell 2004; 116(2): 281-97.
[http://dx.doi.org/10.1016/S0092-8674(04)00045-5] [PMID: 14744438]
[17]
Osada H, Takahashi T. MicroRNAs in biological processes and carcinogenesis. Carcinogenesis 2007; 28(1): 2-12.
[http://dx.doi.org/10.1093/carcin/bgl185] [PMID: 17028302]
[18]
O’Brien J, Hayder H, Zayed Y, Peng C. Overview of microrna biogenesis, mechanisms of actions, and circulation. Front Endocrinol (Lausanne) 2018; 9: 402.
[http://dx.doi.org/10.3389/fendo.2018.00402] [PMID: 30123182]
[19]
Liu J, Zheng M, Tang YL, Liang XH, Yang Q. MicroRNAs, an active and versatile group in cancers. Int J Oral Sci 2011; 3(4): 165-75.
[http://dx.doi.org/10.4248/IJOS11063] [PMID: 22010574]
[20]
Winter J, Jung S, Keller S, Gregory RI, Diederichs S. Many roads to maturity: microRNA biogenesis pathways and their regulation. Nat Cell Biol 2009; 11(3): 228-34.
[http://dx.doi.org/10.1038/ncb0309-228] [PMID: 19255566]
[21]
King VM, Borchert GM. MicroRNA Expression: Protein participants in MicroRNA regulation. Methods Mol Biol 2017; 1617: 27-37.
[http://dx.doi.org/10.1007/978-1-4939-7046-9_2] [PMID: 28540674]
[22]
Ishizuka A, Siomi MC, Siomi H. A drosophila fragile X protein interacts with components of RNAi and ribosomal proteins. Genes Dev 2002; 16(19): 2497-508.
[http://dx.doi.org/10.1101/gad.1022002] [PMID: 12368261]
[23]
Suzuki HI, Yamagata K, Sugimoto K, Iwamoto T, Kato S, Miyazono K. Modulation of microRNA processing by p53. Nature 2009; 460(7254): 529-33.
[http://dx.doi.org/10.1038/nature08199] [PMID: 19626115]
[24]
Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)). Method Methods 2001; 25(4): 402-8.
[http://dx.doi.org/10.1006/meth.2001.1262] [PMID: 11846609]
[25]
García-Cruz R, Camats M, Calin GA, et al. The role of p19 and p21 H-Ras proteins and mutants in miRNA expression in cancer and a Cos-tello syndrome cell model. BMC Med Genet 2015; 16: 46.
[http://dx.doi.org/10.1186/s12881-015-0184-z] [PMID: 26138095]
[26]
Tavazoie SF, Alarcón C, Oskarsson T, et al. Endogenous human microRNAs that suppress breast cancer metastasis. Nature 2008; 451(7175): 147-52.
[http://dx.doi.org/10.1038/nature06487] [PMID: 18185580]
[27]
Jia Z, Zhang Y, Xu Q, Guo W, Guo A. miR-126 suppresses epithelial-to-mesenchymal transition and metastasis by targeting PI3K/AKT/Snail signaling of lung cancer cells. Oncol Lett 2018; 15(5): 7369-75.
[http://dx.doi.org/10.3892/ol.2018.8207] [PMID: 29725450]
[28]
Huo W, Zhang M, Li C, et al. Correlation of microRNA-335 expression level with clinical significance and prognosis in non-small cell lung cancer. Medicine (Baltimore) 2020; 99(34): e21369.
[http://dx.doi.org/10.1097/MD.0000000000021369] [PMID: 32846757]
[29]
Johnson SM, Grosshans H, Shingara J, et al. RAS is regulated by the let-7 microRNA family. Cell 2005; 120(5): 635-47.
[http://dx.doi.org/10.1016/j.cell.2005.01.014] [PMID: 15766527]
[30]
Khodayari N, Mohammed KA, Goldberg EP, Nasreen N. EphrinA1 inhibits malignant mesothelioma tumor growth via let-7 microRNA-mediated repression of the RAS oncogene. Cancer Gene Ther 2011; 18(11): 806-16.
[http://dx.doi.org/10.1038/cgt.2011.50] [PMID: 21869823]
[31]
Yu F, Yao H, Zhu P, et al. let-7 regulates self renewal and tumorigenicity of breast cancer cells. Cell 2007; 131(6): 1109-23.
[http://dx.doi.org/10.1016/j.cell.2007.10.054] [PMID: 18083101]
[32]
Adams BD, Furneaux H, White BA. The micro-ribonucleic acid (miRNA) miR-206 targets the human estrogen receptor-alpha (ERalpha) and represses ERalpha messenger RNA and protein expression in breast cancer cell lines. Mol Endocrinol 2007; 21(5): 1132-47.
[http://dx.doi.org/10.1210/me.2007-0022] [PMID: 17312270]
[33]
Tokumaru S, Suzuki M, Yamada H, Nagino M, Takahashi T. let-7 regulates Dicer expression and constitutes a negative feedback loop. Carcinogenesis 2008; 29(11): 2073-7.
[http://dx.doi.org/10.1093/carcin/bgn187] [PMID: 18700235]
[34]
Pan JY, Sun CC, Bi ZY, et al. miR-206/133b cluster: A weapon against lung cancer? Mol Ther Nucleic Acids 2017; 8: 442-9.
[http://dx.doi.org/10.1016/j.omtn.2017.06.002] [PMID: 28918043]
[35]
Song G, Zhang Y, Wang L. MicroRNA-206 targets notch3, activates apoptosis, and inhibits tumor cell migration and focus formation. J Biol Chem 2009; 284(46): 31921-7.
[http://dx.doi.org/10.1074/jbc.M109.046862] [PMID: 19723635]
[36]
Salzman DW, Shubert-Coleman J, Furneaux H. P68 RNA helicase unwinds the human let-7 microRNA precursor duplex and is required for let-7-directed silencing of gene expression. J Biol Chem 2007; 282(45): 32773-9.
[http://dx.doi.org/10.1074/jbc.M705054200] [PMID: 17724023]
[37]
Emmerth S, Schober H, Gaidatzis D, Roloff T, Jacobeit K, Bühler M. Nuclear retention of fission yeast dicer is a prerequisite for RNAi-mediated heterochromatin assembly. Dev Cell 2010; 18(1): 102-13.
[http://dx.doi.org/10.1016/j.devcel.2009.11.011] [PMID: 20152181]
[38]
Doyle M, Badertscher L, Jaskiewicz L, et al. The double-stranded RNA binding domain of human Dicer functions as a nuclear localization signal. RNA 2013; 19(9): 1238-52.
[http://dx.doi.org/10.1261/rna.039255.113] [PMID: 23882114]
[39]
Burger K, Schlackow M, Potts M, Hester S, Mohammed S, Gullerova M. Nuclear phosphorylated Dicer processes double-stranded RNA in response to DNA damage. J Cell Biol 2017; 216(8): 2373-89.
[http://dx.doi.org/10.1083/jcb.201612131] [PMID: 28642363]
[40]
Burger K, Gullerova M. Nuclear re-localization of Dicer in primary mouse embryonic fibroblast nuclei following DNA damage. PLoS Genet 2018; 14(2): e1007151.
[http://dx.doi.org/10.1371/journal.pgen.1007151] [PMID: 29394246]
[41]
Bronisz A, Rooj AK, Krawczyński K, et al. The nuclear DICER-circular RNA complex drives the deregulation of the glioblastoma cell mi-croRNAome. Sci Adv 2020; 6(51): eabc0221.
[http://dx.doi.org/10.1126/sciadv.abc0221] [PMID: 33328224]

Rights & Permissions Print Cite
Article Metrics
25
1
© 2024 Bentham Science Publishers | Privacy Policy