Abstract
Constituting 5 % of the human genome, microRNAs represent a sizeable class of gene regulators that is predicted to control the expression of at least 60 % of all protein-coding RNAs. Dysregulation of microRNA function results in developmental defects and pathological diseases such as cancers and neurological disorders. Intriguingly, many phenotypes of microRNA deficiencies are subdued in normal condition but manifested apparently upon stress. Here, we outline experimental methods to monitor the level, targets, and activity of microRNAs as the first few steps to characterize how microRNA functions are altered upon stress.
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References
Marsit CJ, Eddy K, Kelsey KT (2006) MicroRNA responses to cellular stress. Cancer Res 66:10843–10848
Leung AK, Sharp PA (2010) MicroRNA functions in stress responses. Mol Cell 40:205–215
Mendell JT, Olson EN (2012) MicroRNAs in stress signaling and human disease. Cell 148:1172–1187
Sturchio E, Colombo T, Boccia P et al (2013) Arsenic exposure triggers a shift in microRNA expression. Sci Total Environ 472C:672–680
Karginov FV, Hannon GJ (2013) Remodeling of Ago2-mRNA interactions upon cellular stress reflects miRNA complementarity and correlates with altered translation rates. Genes Dev 27:1624–1632
Leung AK, Calabrese JM, Sharp PA (2006) Quantitative analysis of Argonaute protein reveals microRNA-dependent localization to stress granules. Proc Natl Acad Sci U S A 103:18125–18130
Wu PH, Isaji M, Carthew RW (2013) Functionally diverse microRNA effector complexes are regulated by extracellular signaling. Mol Cell 52:113–123
Leung AK, Young AG, Bhutkar A et al (2011) Genome-wide identification of Ago2 binding sites from mouse embryonic stem cells with and without mature microRNAs. Nat Struct Mol Biol 18:237–244
Hafner M, Landthaler M, Burger L et al (2010) Transcriptome-wide identification of RNA-binding protein and microRNA target sites by PAR-CLIP. Cell 141:129–141
Kishore S, Jaskiewicz L, Burger L et al (2011) A quantitative analysis of CLIP methods for identifying binding sites of RNA-binding proteins. Nat Methods 8:559–564
Leung AK, Vyas S, Rood JE et al (2011) Poly(ADP-ribose) regulates stress responses and microRNA activity in the cytoplasm. Mol Cell 42:489–499
Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2-ΔΔCT method. Methods 25:402–408
Ingolia NT, Brar GA, Rouskin S et al (2012) The ribosome profiling strategy for monitoring translation in vivo by deep sequencing of ribosome-protected mRNA fragments. Nat Protoc 7:1534–1550
Vourekas A, Mourelatos Z (2014) HITS-CLIP (CLIP-Seq) for mouse Piwi proteins. Methods Mol Biol 1093:73–95
Kincaid RP, Sullivan CS (2012) Virus-encoded microRNAs: an overview and a look to the future. PLoS Pathog 8:e1003018
Huntzinger E, Izaurralde E (2011) Gene silencing by microRNAs: contributions of translational repression and mRNA decay. Nat Rev Genet 12:99–110
Acknowledgement
This work was supported by Department of Defense Breast Cancer Research Program Idea Award #BC101881 to A.K.L.
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Ando, Y., Leung, A.K.L. (2015). Methods for Studying microRNA Functions During Stress. In: Oslowski, C. (eds) Stress Responses. Methods in Molecular Biology, vol 1292. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2522-3_9
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DOI: https://doi.org/10.1007/978-1-4939-2522-3_9
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-2521-6
Online ISBN: 978-1-4939-2522-3
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