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Sequence-specific extracellular microRNAs activate TLR7 and induce cytokine secretion and leukocyte migration

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Abstract

Toll-like receptors (TLRs) can contribute to central nervous system disease pathologies via recognition of microRNAs (miRNAs); however, it remains to be determined which miRNAs are able to activate this signaling. Here we report that numerous miRNAs induced the production of tumor necrosis factor alpha in multiple myeloid cell types, including microglia, and that this effect was abolished in cells deficient in TLR7. Examination of closely related miRNAs that differed in their ability to activate TLR7 resulted in the identification of a motif (UGCUUAU) in miR-20a-5p and specific nucleotides (all the uridines and surprisingly the cytosine as well) in a key area of miR-20a-5p and miR-148b-3p that were vital for the secretion of cytokines via TLR7 stimulation. A 10-nucleotide sequence including this motif was identified to be the shortest single-stranded RNA to signal via TLR7. An miRNA containing this motif induced the secretion of multiple proinflammatory molecules, which was dependent on the phosphoinositide 3-kinase, mitogen-activated protein kinase, and nuclear factor kappa‐light‐chain‐enhancer of activated B cell signaling pathways. Wild-type mice administered miR-20a-5p, which contained this motif, demonstrated increased leukocyte migration. This effect was significantly ameliorated in TLR7-knockout mice, and mice administered miR-20b-5p, in which the motif was mutated, did not exhibit leukocyte migration. We provide a detailed analysis of miRNAs that activate endosomal TLR7 and identify key nucleotide features of a sequence motif recognized by TLR7.

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Acknowledgements

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Funding

This work was supported by the National Institutes of Health [R01 DA040397, R21 MH106422, and P30 MH062261].

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Authors and Affiliations

  1. Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, 42nd and Emile, Omaha, NE, 68198, USA

    Niming Wu, Brenda M. Morsey, Katy M. Emanuel & Howard S. Fox

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  1. Niming Wu
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Contributions

NW and HSF designed experiments; BMM and KME carried out experiments, analyzed experimental results. NW wrote the manuscript, HSF revised the manuscript. All authors approved the final manuscript.

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Correspondence to Howard S. Fox.

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All animal protocols were approved by the Institutional Animal Care and Use Committee of the University of Nebraska Medical Center, and followed the National Institutes of Health guidelines.

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Wu, N., Morsey, B.M., Emanuel, K.M. et al. Sequence-specific extracellular microRNAs activate TLR7 and induce cytokine secretion and leukocyte migration. Mol Cell Biochem 476, 4139–4151 (2021). https://doi.org/10.1007/s11010-021-04220-3

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