Precursor miRNA loop-modulated target regulation
Modulation of mRNA activity is achieved with precursor miRNAs (ta-RNAs). ta-RNAs, primarily pre-miRNAs and pri-miRNAs, including truncated and mutated ta-RNAs, are employed for modulation of mRNA expression where it is found that pri- and pre-miRNA have activity independently of the presence of functional mature miRNAs. Modification of at least one of the stem and loop of the ta-RNAs to enhance binding of the ta-RNA to the target mRNA is employed. The modification may be enhanced complementarity between the ta-RNA and the target mRNA and/or improved thermodynamic efficiency in binding of the ta-RNA to the target.
1. A composition comprising:
a mutated precursor miRNA that hybridizes to a separate target mRNA, wherein said mutated precursor miRNA is from a wild-type precursor miRNA that hybridizes with said separate target mRNA sequence and which has an wild-type terminal loop comprising a wild-type terminal loop sequence, and
wherein said mutated precursor miRNA comprises a stem sequence and a terminal loop, wherein said terminal loop comprises a mutated terminal loop sequence with the same length as said wild-type terminal loop sequence,
wherein at least the first and second nucleotides, and/or the seventh and eighth nucleotides, counting from the 5′ end of said mutated terminal loop sequence, are substituted to enhance or reduce activity of said mutated precursor miRNA in binding and repressing said target mRNA sequence as compared to said wild-type precursor miRNA.
2. The composition of claim 1 , further comprising said separate target mRNA sequence.
3. The composition of claim 1 , wherein said mutated terminal loop sequence comprises at least 8 nucleotides, and wherein said at least 8 nucleotides are numbered from the 5′ end of said loop sequence as follows: position 1, position 2, position 3, position 4, position 5, position 6, position 7, and position 8.
4. The composition of claim 3 , wherein at least said positions 1 and 2 of said mutated terminal loop are substituted compared to said wild-type terminal loop.
5. The composition of claim 3 , wherein at least positions 7 and 8 of said mutated terminal loop are substituted compared to said wild-type terminal loop.