HAIRPIN mRNA ELEMENTS AND METHODS FOR THE REGULATION OF PROTEIN TRANSLATION
AssigneeUniversity of Florida Research Foundation, Inc.
InventorsLing Chen, Arun Srivastava
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Abstract
The present invention provides novel mRNA elements capable of forming hairpin, double-stranded RNA structures independent of other non-coding RNAs. These mRNAs are stably expressed, lack polyadenylation tails, and allow minimal protein translation except when in the presence of specific proteins. Also provided, are compositions and kits comprising the mRNA element, as well as methods for its use in the regulation of protein translation. Advantageously, the disclosed elements represent a novel tool useful in regulating the expression of a wide variety of proteins of interest.
Claims32
A recombinant adeno-associated viral (rAAV) vector comprising at least a first polynucleotide that comprises a promoter operably linked to at least a first nucleic acid segment that encodes at least a first mammalian polypeptide, and wherein the mRNA produced from the vector lacks a polyA tail, but retains an inverted terminal repeat (ITRR) at its 3′-end.
The vector of any preceding claim, wherein an increased translocation of the vector across a mammalian cell membrane is mediated by one or more endogenous or exogenously-provided cellular proteins, regulatory elements, siRNAs, helper plasmids, or any combination thereof.
The vector of any preceding claim, further comprising a reporter gene, such as that encoding GFP.
The vector of any preceding claim, wherein expression of the encoded polypeptide is regulated by the presence of a helper plasmid that encodes one or more cellular or viral proteins, or a combination thereof.
The vector of any preceding claim, wherein the promoter is an inducible or a regulatable promoter.
The rAAV vector of any preceding claim, wherein the vector is a self-complementary rAAV (scAAV).
The rAAV vector according to any preceding claim, further comprising at least a first enhancer operably linked to the at least a first nucleic segment.
The rAAV vector according to any preceding claim, further comprising at least a first mammalian intron sequence operably linked to the at least a first nucleic segment.
The rAAV vector according to any preceding claim, wherein the at least a first mammalian polypeptide is of human, non-human primate, murine, feline, canine, porcine, ovine, bovine, equine, epine, caprine, or lupine origin.
The rAAV vector according to any preceding claim, wherein the at least a first nucleic acid segment encodes a biologically-active human polypeptide.
The rAAV vector according to any preceding claim, wherein the rAAV vector is a recombinant adeno-associated virus serotype 1 (rAAV1) vector, a recombinant adeno-associated virus serotype 2 (rAAV2) vector, recombinant adeno-associated virus serotype 3 (rAAV3) vector, a recombinant adeno-associated virus serotype 4 (rAAV4) vector, a recombinant adeno-associated virus serotype 5 (rAAV5) vector, a recombinant adeno-associated virus serotype 6 (rAAV6) vector, a recombinant adeno-associated virus serotype 7 (rAAV7) vector, a recombinant adeno-associated virus serotype 8 (rAAV8) vector, a recombinant adeno-associated virus serotype 9 (rAAV9) vector, a recombinant adeno-associated virus serotype 10 (rAAV10) vector, a recombinant adeno-associated virus serotype 11 (rAAV11) vector, or a recombinant adeno-associated virus serotype 12 (rAAV12) vector.
The rAAV vector according to any preceding claim, comprised within an infectious adeno-associated viral particle, virion, or a plurality of infectious AAV particles.
Dependencies are inferred from the claim text.