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The regulatory activity of microRNA* species has substantial influence on microRNA and 3′ UTR evolution

Nature Structural & Molecular Biology volume 15, pages 354–363 (2008)Cite this article

Abstract

During microRNA (miRNA) biogenesis, one strand of a ∼21–22-nucleotide RNA duplex is preferentially selected for entry into a silencing complex. The other strand, known as the miRNA* species, has typically been assumed to be a carrier strand. Here we show that, although Drosophila melanogaster miRNA* species are less abundant than their partners, they are often present at physiologically relevant levels and can associate with Argonaute proteins. Comparative genomic analyses revealed that >40% of miRNA* sequences resist nucleotide divergence across Drosophilid evolution, and at least half of these well-conserved miRNA* species select for conserved 3′ untranslated region seed matches well above background noise. Finally, we validated the inhibitory activity of miRNA* species in both cultured cells and transgenic animals. These data broaden the reach of the miRNA regulatory network and suggest an important mechanism that diversifies miRNA function during evolution.

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Figure 1: Both miRNA and miRNA* species can be detected in total RNA.
Figure 2: Highly conserved miRNA* species accumulate to higher relative levels at steady state.
Figure 3: miRNA and miRNA* species can be co-immunoprecipitated with AGO1.
Figure 4: Sensor assays in cultured cells and transgenic animals validate the regulatory activity of miRNA* species.
Figure 5: Bioinformatic evidence for the endogenous usage of both miRNAs and miRNA* species as regulatory RNAs.
Figure 6: Endogenous relevance of miRNA*-mediated repression.
Figure 7: The inherent dual-regulatory capability of miRNA hairpins may influence miRNA evolution.

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Acknowledgements

We thank B. Tam (University of California, Davis) for helping to construct transgenic sensors. We thank J.G. Ruby and D. Bartel (Howard Hughes Medical Institute and Whitehead Institute) for sharing their initial analysis of Drosophila miRNAs; S. Crews (University of North Carolina) for the Abrupt antibody; the University of California, Santa Cruz genome center, Agencourt, and the Baylor College of Medicine for Drosophila genome sequences, assemblies and alignments; and J. Major (Sloan-Kettering Institute) for software and technical support. K.O. was supported by a grant from the Charles Revson Foundation. E.C.L. was supported by grants from the Leukemia and Lymphoma Society, the Burroughs Wellcome Foundation, the V Foundation for Cancer Research, the Sidney Kimmel Cancer Foundation and the US National Institutes of Health (GM083300).

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Authors and Affiliations

  1. Department of Developmental Biology, Sloan-Kettering Institute, 521 Rockefeller Research Laboratories, 1275 York Ave, Box 252, New York, 10065, New York, USA

    Katsutomo Okamura, Michael D Phillips, David M Tyler, Hong Duan, Yu-ting Chou & Eric C Lai

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  1. Katsutomo Okamura
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  2. Michael D Phillips
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  4. Hong Duan
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Contributions

K.O. performed the northern blots, IP tests and luciferase sensor assays; M.D.P. performed the computational analysis; D.M.T. performed imaginal disc stainings; H.D. and Y.-t.C. prepared sensor constructs; E.C.L. wrote the manuscript.

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Correspondence to Eric C Lai.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–4 and Supplementary Methods (PDF 7803 kb)

Supplementary Table 1

miRNA/miRNA* clone counts and ratios across Drosophila development. (XLS 125 kb)

Supplementary Table 2

Segregated clone data of 26 genes that generated at least 50 reads in all 4 embryonic time windows. (XLS 38 kb)

Supplementary Table 3

Target signal noise and P-values for the 65 highly conserved (HC) miRNA and miRNA* seeds. (XLS 24 kb)

Supplementary Table 4

Target signal noise and P-values for the 20 poorly conserved (PC) miRNA and miRNA* seeds. (XLS 19 kb)

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Okamura, K., Phillips, M., Tyler, D. et al. The regulatory activity of microRNA* species has substantial influence on microRNA and 3′ UTR evolution. Nat Struct Mol Biol 15, 354–363 (2008). https://doi.org/10.1038/nsmb.1409

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