Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Experiment Videos

A novel assay for viral microRNA function identifies a single nucleotide polymorphism that affects Drosha processing.

Eva Gottwein1, Xuezhong Cai, Bryan R Cullen

  • 1Department of Molecular Genetics and Microbiology, Box 3025, Duke University Medical Center, Durham, NC 27710, USA.

Journal of Virology
|May 16, 2006
PubMed
Summary

This study developed a new assay to demonstrate the function of microRNAs (miRNAs) from Kaposi's sarcoma-associated herpesvirus (KSHV). The assay confirmed viral miRNA activity and identified a polymorphism affecting miR-K5 processing.

Related Concept Videos

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

Cytotoxicity of activator expression in CRISPR-based transcriptional activation systems.

Nature communications·2025
Same author

KSHV-encoded vIRF3 Cooperates with Cellular IRF4 to Drive Super-Enhancer Activity through Complex DNA Elements.

bioRxiv : the preprint server for biology·2025
Same author

Efficacious genome editing in infant mice with glycogen storage disease type Ia.

JCI insight·2025
Same author

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

Nucleic acids research·2024
Same author

Cytotoxicity of Activator Expression in CRISPR-based Transcriptional Activation Systems.

bioRxiv : the preprint server for biology·2024
Same author

Author Correction: RNA conformational propensities determine cellular activity.

Nature·2023

Area of Science:

  • Virology
  • Molecular Biology
  • Genetics

Background:

  • MicroRNAs (miRNAs) are key regulators of gene expression.
  • Kaposi's sarcoma-associated herpesvirus (KSHV) encodes 11 miRNAs, but their function in target gene inhibition was unproven.
  • Assessing viral miRNA function requires sensitive and specific assays.

Purpose of the Study:

  • To develop and validate a novel assay for measuring viral miRNA function.
  • To demonstrate the posttranscriptional inhibitory activity of KSHV-encoded miRNAs.
  • To investigate the molecular basis for differential activity of a specific viral miRNA (miR-K5).

Main Methods:

  • Development of a lentiviral indicator vector system with miRNA target sites in the Renilla luciferase 3' UTR.
  • Co-transduction of indicator and control vectors in KSHV-infected and uninfected cells.

Related Experiment Videos

  • Sequence analysis of viral genomes and in vitro/in vivo Drosha processing assays.

    • Main Results:

    • The novel assay successfully demonstrated the inhibitory function of KSHV miRNAs in infected cells, reducing Renilla luciferase expression up to 10-fold.
    • A single nucleotide polymorphism in the miR-K5 precursor stem-loop was identified in BCBL-1 cells.
    • This polymorphism diminished Drosha processing of pri-miR-K5, leading to reduced mature miR-K5 levels and function.

    Conclusions:

    • The developed lentiviral assay is effective for assessing viral miRNA function.
    • Naturally occurring sequence variations in miRNA precursors can significantly impact miRNA processing and activity.
    • This study provides the first evidence of a polymorphism in an miRNA precursor affecting Drosha processing and mature miRNA levels.