Synthetic Design of Asymmetric miRNA with an Engineered 3′ Overhang to Improve Strand Selection

Sandeep Kadekar, Ganesh N. Nawale, Kira Karlsson, Cecilia Ålander, Oommen P. Oommen, Oommen P. Varghese

    Research output: Contribution to journalArticleScientificpeer-review

    6 Citations (Scopus)
    7 Downloads (Pure)

    Abstract

    We developed a novel miRNA design that significantly improves strand selection within the RISC complex by engineering the 3′ end by adding extra nucleotides. Addition of seven nucleotides at the 3′ ends of the miR or miR* strand resulted in a thermodynamic asymmetry at either of the two ends, which resulted in selective RISC recruitment, as demonstrated by a stem-loop PCR experiment. Such selective recruitment was also corroborated at the protein level by western blot analysis. To investigate the functional effect because of selective recruitment, we performed apoptosis and metastasis studies using human colon carcinoma cells (HCT116)and human osteosarcoma cells (MG63). These experiments indicated that recruitment of the miR strand is responsible for inducing apoptosis and inhibiting the invasiveness of cancer cells. Recruitment of the miR* strand, on the other hand, had the opposite effect. To the best of our knowledge, our strand engineering strategy is the first report of improved strand selection of a desired miRNA strand by RISC without using any chemical modifications or mismatches. We believe that such structural modifications of miR34a could mitigate some of the off-target effects of miRNA therapy and would also allow a better understanding of sequence-specific gene regulation. Such a design could also be adapted to other miRNAs to enhance their therapeutic potential.

    Original languageEnglish
    Pages (from-to)597-604
    Number of pages8
    JournalMolecular Therapy - Nucleic Acids
    Volume16
    DOIs
    Publication statusPublished - 7 Jun 2019
    Publication typeA1 Journal article-refereed

    Keywords

    • anticancer therapy
    • miR34a
    • miRNA
    • RNAi
    • strand selection

    Publication forum classification

    • Publication forum level 1

    ASJC Scopus subject areas

    • Molecular Medicine
    • Drug Discovery

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