title: Analyzing mRNA expression identifies Smad3 as a microRNA-140 target regulated only at protein level.
abstract: mRNA profiling is routinely used to identify microRNA targets, however, this high-throughput technology is not suitable for identifying targets regulated only at protein level. Here, we have developed and validated a novel methodology based on computational analysis of promoter sequences combined with mRNA microarray experiments to reveal transcription factors that are direct microRNA targets at the protein level. Using this approach we identified Smad3, a key transcription factor in the TGFbeta signaling pathway, as a direct miR-140 target. We showed that miR-140 suppressed the TGFbeta pathway through repression of Smad3 and that TGFbeta suppressed the accumulation of miR-140 forming a double negative feedback loop. Our findings establish a valid strategy for the discovery of microRNA targets regulated only at protein level, and we propose that additional targets could be identified by re-analysis of existing microarray datasets.
authors.affiliation: School of Computing Sciences, University of East Anglia, Norwich, NR4 7TJ, United Kingdom.
authors.forename: Helio
authors.initials: H
authors.lastname: Pais
authors.forename: Francisco Esteban
authors.initials: FE
authors.lastname: Nicolas
authors.forename: Surinder M
authors.initials: SM
authors.lastname: Soond
authors.forename: Tracey E
authors.initials: TE
authors.lastname: Swingler
authors.forename: Ian M
authors.initials: IM
authors.lastname: Clark
authors.forename: Andrew
authors.initials: A
authors.lastname: Chantry
authors.forename: Vincent
authors.initials: V
authors.lastname: Moulton
authors.forename: Tamas
authors.initials: T
authors.lastname: Dalmay
mesh_terms: D000818:Animals; D002460:Cell Line; D019295:Computational Biology / Q000379:methods*; D051379:Mice; D035683:MicroRNAs / Q000378:metabolism*; D020411:Oligonucleotide Array Sequence Analysis; D011401:Promoter Regions, Genetic; D051900:Smad3 Protein / Q000378:metabolism*; D016212:Transforming Growth Factor beta / Q000378:metabolism
chemical_list: C547828:MIRN140 microRNA, mouse; D035683:MicroRNAs; D051900:Smad3 Protein; C497791:Smad3 protein, mouse; D016212:Transforming Growth Factor beta