RNA-Binding Protein Motifs Predict MicroRNA Secretion and Cellular Retention in Hypothalamic and Other Cell Types

Overview

Journal Biomedicines

Specialty Biomedical Engineering

Date 2024 Apr 27

PMID 38672211

Authors

Wenyuan He

Denise D Belsham

Affiliations

Soon will be listed here.

Abstract

Cellular microRNAs (miRNAs) can be selectively secreted or retained, adding another layer to their critical role in regulating human health and disease. To date, select RNA-binding proteins (RBPs) have been proposed to be a mechanism underlying miRNA localization, but the overall relevance of RBPs in systematic miRNA sorting remains unclear. This study profiles intracellular and small extracellular vesicles' (sEVs) miRNAs in NPY-expressing hypothalamic neurons. These findings were corroborated by the publicly available sEV and intracellular miRNA profiles of white and brown adipocytes, endothelium, liver, and muscle from various databases. Using experimentally determined binding motifs of 93 RBPs, our enrichment analysis revealed that sEV-originating miRNAs contained significantly different RBP motifs than those of intracellularly retained miRNAs. Multiple RBP motifs were shared across cell types; for instance, RBM4 and SAMD4 are significantly enriched in neurons, hepatocytes, skeletal muscle, and endothelial cells. Homologs of both proteins physically interact with Argonaute1/2 proteins, suggesting that they play a role in miRNA sorting. Machine learning modelling also demonstrates that significantly enriched RBP motifs could predict cell-specific preferential miRNA sorting. Non-optimized machine learning modeling of the motifs using Random Forest and Naive Bayes in all cell types except WAT achieved an area under the receiver operating characteristic (ROC) curve of 0.77-0.84, indicating a high predictive accuracy. Given that the RBP motifs have a significant predictive power, these results underscore the critical role that RBPs play in miRNA sorting within mammalian cells and reinforce the importance of miRNA sequencing in preferential localization. For the future development of small RNA therapeutics, considering these RBP-RNA interactions could be crucial to maximize delivery effectiveness and minimize off-target effects.

References

Karagkouni D, Paraskevopoulou M, Chatzopoulos S, Vlachos I, Tastsoglou S, Kanellos I . DIANA-TarBase v8: a decade-long collection of experimentally supported miRNA-gene interactions. Nucleic Acids Res. 2017; 46(D1):D239-D245. PMC: 5753203. DOI: 10.1093/nar/gkx1141. View

Chen Y, Zhao Y, Yin Y, Jia X, Mao L . Mechanism of cargo sorting into small extracellular vesicles. Bioengineered. 2021; 12(1):8186-8201. PMC: 8806638. DOI: 10.1080/21655979.2021.1977767. View

Frank E, Hall M, Trigg L, Holmes G, Witten I . Data mining in bioinformatics using Weka. Bioinformatics. 2004; 20(15):2479-81. DOI: 10.1093/bioinformatics/bth261. View

Xu D, Di K, Fan B, Wu J, Gu X, Sun Y . MicroRNAs in extracellular vesicles: Sorting mechanisms, diagnostic value, isolation, and detection technology. Front Bioeng Biotechnol. 2022; 10:948959. PMC: 9618890. DOI: 10.3389/fbioe.2022.948959. View

Wei Z, Batagov A, Schinelli S, Wang J, Wang Y, El Fatimy R . Coding and noncoding landscape of extracellular RNA released by human glioma stem cells. Nat Commun. 2017; 8(1):1145. PMC: 5658400. DOI: 10.1038/s41467-017-01196-x. View

Krol J, Busskamp V, Markiewicz I, Stadler M, Ribi S, Richter J . Characterizing light-regulated retinal microRNAs reveals rapid turnover as a common property of neuronal microRNAs. Cell. 2010; 141(4):618-31. DOI: 10.1016/j.cell.2010.03.039. View

Santangelo L, Giurato G, Cicchini C, Montaldo C, Mancone C, Tarallo R . The RNA-Binding Protein SYNCRIP Is a Component of the Hepatocyte Exosomal Machinery Controlling MicroRNA Sorting. Cell Rep. 2016; 17(3):799-808. DOI: 10.1016/j.celrep.2016.09.031. View

Crooke S, Vickers T, Liang X . Phosphorothioate modified oligonucleotide-protein interactions. Nucleic Acids Res. 2020; 48(10):5235-5253. PMC: 7261153. DOI: 10.1093/nar/gkaa299. View

Mori M, Ludwig R, Garcia-Martin R, Brandao B, Kahn C . Extracellular miRNAs: From Biomarkers to Mediators of Physiology and Disease. Cell Metab. 2019; 30(4):656-673. PMC: 6774861. DOI: 10.1016/j.cmet.2019.07.011. View

10.

McIlwraith E, Belsham D . Palmitate alters miRNA content of small extracellular vesicles secreted from NPY/AgRP-expressing hypothalamic neurons. Brain Res. 2023; 1810:148367. DOI: 10.1016/j.brainres.2023.148367. View

11.

Friedrich M, Aigner A . Therapeutic siRNA: State-of-the-Art and Future Perspectives. BioDrugs. 2022; 36(5):549-571. PMC: 9396607. DOI: 10.1007/s40259-022-00549-3. View

12.

Thomou T, Mori M, Dreyfuss J, Konishi M, Sakaguchi M, Wolfrum C . Adipose-derived circulating miRNAs regulate gene expression in other tissues. Nature. 2017; 542(7642):450-455. PMC: 5330251. DOI: 10.1038/nature21365. View

13.

Isaac R, Reis F, Ying W, Olefsky J . Exosomes as mediators of intercellular crosstalk in metabolism. Cell Metab. 2021; 33(9):1744-1762. PMC: 8428804. DOI: 10.1016/j.cmet.2021.08.006. View

14.

Mayer C, Belsham D . Insulin directly regulates NPY and AgRP gene expression via the MAPK MEK/ERK signal transduction pathway in mHypoE-46 hypothalamic neurons. Mol Cell Endocrinol. 2009; 307(1-2):99-108. DOI: 10.1016/j.mce.2009.02.031. View

15.

La Rocca G, Olejniczak S, Gonzalez A, Briskin D, Vidigal J, Spraggon L . In vivo, Argonaute-bound microRNAs exist predominantly in a reservoir of low molecular weight complexes not associated with mRNA. Proc Natl Acad Sci U S A. 2015; 112(3):767-72. PMC: 4311832. DOI: 10.1073/pnas.1424217112. View

16.

Bailey T, Johnson J, Grant C, Noble W . The MEME Suite. Nucleic Acids Res. 2015; 43(W1):W39-49. PMC: 4489269. DOI: 10.1093/nar/gkv416. View

17.

Garcia-Martin R, Wang G, Brandao B, Zanotto T, Shah S, Patel S . MicroRNA sequence codes for small extracellular vesicle release and cellular retention. Nature. 2021; 601(7893):446-451. PMC: 9035265. DOI: 10.1038/s41586-021-04234-3. View

18.

Villarroya-Beltri C, Gutierrez-Vazquez C, Sanchez-Cabo F, Perez-Hernandez D, Vazquez J, Martin-Cofreces N . Sumoylated hnRNPA2B1 controls the sorting of miRNAs into exosomes through binding to specific motifs. Nat Commun. 2013; 4:2980. PMC: 3905700. DOI: 10.1038/ncomms3980. View

19.

Tosar J, Gambaro F, Sanguinetti J, Bonilla B, Witwer K, Cayota A . Assessment of small RNA sorting into different extracellular fractions revealed by high-throughput sequencing of breast cell lines. Nucleic Acids Res. 2015; 43(11):5601-16. PMC: 4477662. DOI: 10.1093/nar/gkv432. View

20.

Ray D, Kazan H, Cook K, Weirauch M, Najafabadi H, Li X . A compendium of RNA-binding motifs for decoding gene regulation. Nature. 2013; 499(7457):172-7. PMC: 3929597. DOI: 10.1038/nature12311. View