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Human let-7a miRNA blocks protein production on actively translating polyribosomes

Nature Structural & Molecular Biology volume 13, pages 1108–1114 (2006)Cite this article

Abstract

MicroRNAs (miRNAs) regulate gene expression at a post-transcriptional level through base-pairing to 3′ untranslated regions (UTRs) of messenger RNAs. The mechanism by which human let-7a miRNA regulates mRNA translation was examined in HeLa cells expressing reporter mRNAs containing the Caenorhabditis elegans lin-41 3′ UTR. let-7a miRNA strongly repressed translation, yet the majority of control and lin-41–bearing RNAs sedimented with polyribosomes in sucrose gradients; these polyribosomes, together with let-7a miRNA and the miRISC protein AGO, were released from those structures by puromycin. RNA containing the lin-41 3′ UTR and an iron response element in the 5′ UTR sedimented with polysomes when cells were incubated with iron, but showed ribosome run-off when the iron was chelated. These data indicate that let-7a miRNA inhibits actively translating polyribosomes. Nascent polypeptide coimmunoprecipitation experiments further suggest that let-7a miRNA interferes with the accumulation of growing polypeptides.

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Figure 1: let-7a miRNA–mediated translational repression in HeLa cells.
Figure 2: lin-41–containing mRNA and let-7a miRNA are associated with translating polyribosomes.
Figure 3: Block of translation initiation results in ribosome run-off from repressed mRNAs.
Figure 4: Polyribosomes are not coimmunoprecipitated when under miRNA control.

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Acknowledgements

We thank C. Mello for discussion and Z. Mourelatos (University of Pennsylvania) for kindly providing the monoclonal antibody to AGO, 28A. S.N. was supported by a fellowship of the Max Planck Society and by European Molecular Biology Organization fellowship ALTF 995-2004. M.J.S. was a Canadian Institutes of Health Research postdoctoral fellow. This work was supported by grants from the US National Institutes of Health (GM46779 and HD37267).

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  1. Martin J Simard

    Present address: Laval University Cancer Research Center, Quebec City, Quebec, Canada

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  1. Program in Molecular Medicine, University of Massachusetts Medical School, 373 Plantation St., Suite 204, Worcester, 01605, Massachusetts, USA

    Stephanie Nottrott, Martin J Simard & Joel D Richter

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Contributions

S.N. performed all the experiments, M.J.S. cloned the lin-41 3′ UTR and S.N. and J.D.R. analyzed the data and wrote the manuscript.

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Correspondence to Joel D Richter.

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The authors declare no competing financial interests.

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Nottrott, S., Simard, M. & Richter, J. Human let-7a miRNA blocks protein production on actively translating polyribosomes. Nat Struct Mol Biol 13, 1108–1114 (2006). https://doi.org/10.1038/nsmb1173

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