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Nucleic acid nanocapsules: packaging mRNA into the vault particle
Author(s) -
Pupols Melody,
Kickhoefer Valerie A.,
Rome Leonard H.
Publication year - 2010
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.24.1_supplement.886.1
Subject(s) - nucleic acid , messenger rna , rna , microbiology and biotechnology , chemistry , lipofectamine , fusion protein , translation (biology) , green fluorescent protein , vault (architecture) , nanocapsules , biophysics , biology , biochemistry , nanotechnology , recombinant dna , materials science , gene , vector (molecular biology) , structural engineering , nanoparticle , engineering
Our goal is to package large (~1000nt) mRNAs inside the vault particle and subsequently use these nucleic acid nanocapsules to deliver mRNA for translation in vivo . Vaults are ubiquitous intracellular components found in most eukaryotes, including humans. They are large (13MDa) ribonucleoprotein particles (72.5 × 41 nm) and are present at 10 4 –10 5 particles per cell. Their interior cavity (5×10 7 Å 3 ) is large enough to encapsulate hundreds of proteins, giving them the potential to be used as biocompatible delivery vehicles.We have created a fusion protein that both targets the interior of vaults (mINT domain) and binds RNA (bacteriophage MS2 dimer). We demonstrate that this fusion protein can bind the untranslated stem loop binding sites of an in vitro synthesized and capped mRNA which encodes GFP. Furthermore, the binding of the fusion protein does not inhibit translation of GFP in vitro. Transfection of the mRNA into mammalian cells with Lipofectamine shows it is functional in vivo , with a minimum of 5ng required for GFP detection. In conclusion, these studies suggest that vaults could be engineered as a novel delivery vehicle for larger RNA transcripts. Funded by the NIH National Institute of Biomedical Imaging and Bioengineering (R01 EB004553) and the University of California Discovery Grant in partnership with Corporate Sponsor Abraxis Bioscience Inc. (BIO07‐10671)