Retargeting target-directed microRNA-decay sites to highly expressed viral or cellular miRNAs

Area of Science:

• Molecular Biology
• Gene Regulation
• Virology

Background:

• MicroRNAs (miRNAs) are key gene expression regulators.
• Target-directed miRNA decay (TDMD) degrades miRNAs via specific base-pairing.
• Existing TDMD sites can be repurposed for miRNA inhibition.

Purpose of the Study:

• To explore retargeting known TDMD sites for inhibiting heterologous miRNAs, including viral miRNAs.
• To investigate the efficacy of Cyrano long non-coding RNA-based inhibitors against miR-K11, a Kaposi's Sarcoma-associated herpesvirus (KSHV) miRNA.
• To assess the impact of miR-K11 inhibition on KSHV-transformed cells and the role of ZSWIM8 in TDMD.

Main Methods:

• Designed and tested miRNA inhibitors based on the Cyrano TDMD site architecture.
• Evaluated inhibitor specificity for viral miR-K11 versus cellular miR-155.
• Delivered inhibitors via lentivirus into KSHV-transformed primary effusion lymphoma (PEL) cells.
• Assessed the effect of ZSWIM8 inactivation on inhibitor efficacy.

Main Results:

• Cyrano-like inhibitors effectively inhibited heterologous miRNAs, outperforming other retargeted sites.
• Inhibitors demonstrated specificity for miR-K11 over miR-155 and vice versa.
• miR-K11 inhibition reduced PEL cell viability, indicating miR-K11's role in KSHV-driven survival.
• ZSWIM8 inactivation did not significantly impair inhibition by Cyrano-based miRNA inhibitors.

Conclusions:

• Natural TDMD sites can be successfully retargeted to inhibit highly expressed viral or cellular miRNAs.
• Cyrano-based inhibitors offer a promising strategy for specific miRNA targeting.
• This study defines key features of effective encoded miRNA inhibitors and highlights miR-K11's role in PEL cell survival.