Isolation of microRNA targets by miRNP immunopurification
Abstract
microRNAs (miRNAs) serve as post-transcriptional regulators of gene expression, by guiding effector complexes (miRNPs) to target RNAs. Although considerable progress has been made in computational methods to identify miRNA targets, only a relatively limited assessment of their ability to function in vivo has been reported. Here we describe an alternative approach to miRNA target identification based on a biochemical method for purifying miRNP complexes with associated miRNAs and bound mRNA targets. Microarray analysis revealed a high degree of enrichment for miRNA complementary sites in the 3′UTRs of the miRNP-associated mRNAs. mRNAs specifically associated with an individual miRNA were identified by comparing the miRNP-associated mRNAs from wild-type flies and mutant flies lacking miR-1, and their regulation by the miRNA was validated. This approach provides a means to identify functional miRNA targets based on their physical interaction in vivo.
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AGO immunoprecipitation has been successfully used to isolate miRNAs and their mRNA targets.
“AGO immunoprecipitation has been successfully used to isolate miRNAs and their mRNA targets (Easow et al., 2007, Beitzinger et al., 2007, Hendrickson et al., 2008, Zhang et al., 2007, Hammell et al., 2008, Karginov et al., 2007).”
From Cell-Type-Based Analysis of MicroRNA Profiles in the Mouse Brain (2012) · cites this paper as Easow et al., 2007
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p16INK4a Translation Suppressed by miR-24
Lal, A., Kim, H. H., Abdelmohsen, K., Kuwano, Y., Pullmann, R., Srikantan, S., Subrahmanyam, R., Martindale, J. L., Yang, X., Ahmed, F., et al. (2008) · PLoS ONE · doi:10.1371/journal.pone.0001864 · Cited by 268 · see citing works
“Evidence that miR-24 interacted with the p16 mRNA was then obtained using a method previously reported to study the functional effects of miRNAs on target mRNAs [29].”
General Principals of miRNA Biogenesis and Regulation in the Brain
O'Carroll, D., & Schaefer, A. (2013) · Neuropsychopharmacology · doi:10.1038/npp.2012.87 · Cited by 205 · see citing works
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- Authors
- Easow, G., Teleman, A. A., & Cohen, S. M.
- Venue
- RNA
- Publisher
- Cold Spring Harbor Laboratory
- Published
- 2007-06-25
- Volume
- 13
- Issue
- 8
- Pages
- 1198-1204
- ISSN
- 1355-8382
- 1469-9001
- Cited by
- 252
- References
- 3
- Language
- en
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