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Integrated analysis of directly captured microRNA targets reveals the impact of microRNAs on mammalian transcriptome.

ABSTRACT: MicroRNA (miRNA)-mediated regulation is widespread, relatively mild but functionally important. It remains challenging to unequivocally identify miRNA targeted RNAs at a genomic scale and determine how changes in miRNA levels affect the transcriptome. Here, we captured individual miRNAs and their targeted RNA sites in wild-type, miR-200 family knockout and induced epithelial cells. We detected 1797 miRNAs interacting with 13,830 transcripts at 616,127 sites by sequencing 1,230,019 unique miRNA:RNA chimeras. Although mRNA sites that are bound by miRNAs and contain matches to seed sequences confer the strongest regulation, ?40%-60% of miRNA bound regions do not contain seed matches. Different miRNAs have different preferences to seed matches and 3' end base-pairing. For individual miRNAs, the effectiveness of mRNA regulation is highly correlated with the number of captured miRNA:mRNA chimeras. Notably, elevated miR-200 expression robustly represses existing targets with little impact on newly recognized targets. Global analysis of directly captured mRNA targets reveals pathways that are involved in cancer and cell adhesion and signaling pathways that are highly regulated by many different miRNAs in epithelial cells. Comparison between experimentally captured and TargetScan predicted targets indicates that our approach is more effective in identifying bona fide targets by reducing false positive and negative predictions. This study reveals the global binding landscape and impact of miRNAs on the mammalian transcriptome.

SUBMITTER: Bjerke GA

PROVIDER: S-EPMC7025510 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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Integrated analysis of directly captured microRNA targets reveals the impact of microRNAs on mammalian transcriptome.

Bjerke Glen A GA Yi Rui R

RNA (New York, N.Y.) 20200103 3

MicroRNA (miRNA)-mediated regulation is widespread, relatively mild but functionally important. It remains challenging to unequivocally identify miRNA targeted RNAs at a genomic scale and determine how changes in miRNA levels affect the transcriptome. Here, we captured individual miRNAs and their targeted RNA sites in wild-type, miR-200 family knockout and induced epithelial cells. We detected 1797 miRNAs interacting with 13,830 transcripts at 616,127 sites by sequencing 1,230,019 unique miRNA:R ...[more]

PMID: 31900330

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Project description:We have employed a novel approach for the identification of functionally important microRNA (miRNA)-target interactions using integrated miRNA, transcriptome and proteome profiles with advanced in silico analysis. By looking at both the transcript and protein levels of expression, a thorough coverage of miRNA regulation was obtained. Microdissected auditory and vestibular sensory epithelia were used as the model system, thus being the first time such a comparison was carried out in a neuroepithelial system. Moreover, this is one of only a few studies employing proteome screening for the identification of miRNA targets. Notably, this approach can be employed for the study of other tissues and organs. We detected the expression of 157 miRNAs in the inner ear sensory epithelia, with 53 miRNAs differentially expressed between the cochlea and vestibule. By searching for enrichment and depletion of miRNA targets in the transcript and protein datasets with a reciprocal or similar expression, respectively, as the regulatory miRNA, we identified functionally important miRNAs. Finally, the interaction between miR-135b and PSIP1-P75, a transcriptional coacitvator previously unknown in the inner ear, was identified and validated experimentally. We suggest that miR-135b may serve as a cellular effector, involved in regulating some of the differences between the cochlear and vestibular hair cells. We investigated the mRNA expression profile of the cochlear and vestibular sensory epithelia from inner ears of postnatal day 2 mice using the Affymetrix GeneChip® 430 2Mouse Genome array. Cochlear and vestibular sensory epithelia were dissected from wild type C3H mice and collected separately. The vestibular epithelia consisted of the saccule, utricle and the lateral and anterior cristae. Both the cochlear and vestibular sensory epithelia were dissected with their underlying mesenchyma. Altogether three pools, three biological replicates, of each tissue type were collected consisting of cochlear or vestibular sensory epithelia dissected from 10 to 12 inner ears.

Dataset Information

Integrated analysis of directly captured microRNA targets reveals the impact of microRNAs on mammalian transcriptome.

Publications

Integrated analysis of directly captured microRNA targets reveals the impact of microRNAs on mammalian transcriptome.

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