Synthesis and delivery of short, noncoding RNA by B lymphocytes

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 110; no. 50; pp. 20182 - 20187
• Main Authors: Almanza, Gonzalo, Anufreichik, Veronika, Rodvold, Jeffrey J., Chiu, Kevin T., DeLaney, Alexandra, Akers, Johnny C., Chen, Clark C., Zanetti, Maurizio
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences 10.12.2013; NATIONAL ACADEMY OF SCIENCES; National Acad Sciences
• Subjects: Animals; Antibodies - immunology; Antigen presenting cells; Antigens; B lymphocytes; B-Lymphocytes - metabolism; Biological Sciences; Biosynthesis; Cell lines; Cross priming; Deoxyribonucleic acid; DNA; Flow Cytometry; Gene expression; Gene Expression Regulation - genetics; Gene Targeting - methods; Humans; Immunotherapy - methods; Lymphocytes; Mice; Mice, Inbred C57BL; MicroRNA; MicroRNAs; MicroRNAs - genetics; MicroRNAs - metabolism; Microscopy, Fluorescence; Molecules; Oligonucleotides - genetics; Ova; Plasmids; Plasmids - genetics; Real-Time Polymerase Chain Reaction; RNA, Small Untranslated - biosynthesis; RNA, Small Untranslated - metabolism; T lymphocytes; Transfection; immunotherapy; microvesicles
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.1311145110

Evolutionarily conserved short (20–30 nucleotides) noncoding RNAs (microRNAs) are powerful regulators of gene expression in a variety of physiological and pathological processes. As such, means to efficiently modulate microRNA function constitute an important therapeutic opportunity. Here we demonstrate that primary B lymphocytes can be genetically programmed with nonviral plasmid DNA for the biogenesis and delivery of antisense sequences (anti-microRNA) against microRNA-150 (miR-150). Within 18 h of transfection with an anti-miR-150 construct, primary B lymphocytes secrete ∼3,000 copies of anti-miR-150 molecules per cell. Anti-miR-150 molecules released by B lymphocytes were internalized by CD8 T lymphocytes during cross-priming in vitro and in vivo, resulting in marked down-regulation of endogenous miR-150. However, such internalization was not observed in the absence of cross-priming. These results suggest that shuttling anti-miR-150 molecules from B lymphocytes to T cells requires the activation of receiver T cells via the antigen receptor. Finally, anti-miR-150 synthesized in B cells were secreted both as free and extracellular vesicle-associated fractions, but only extracellular vesicle-associated anti-miR-150 were apparently taken up by CD8 T cells. Collectively, these data indicate that primary B lymphocytes represent an efficient platform for the synthesis and delivery of short, noncoding RNA, paving the way for an approach to immunogenomic therapies.