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 method for identifying the role of a loop sequence of a precursor miRNA among a plurality of isoforms differing in their loop sequences, said method comprising:
substituting one loop from one isoform with an equivalent loop from a different isoform to produce a mutated precursor miRNA;
introducing said one isoform in a first cell and said mutated precursor miRNA into a second cell;
determining the effect of the one isoform and said mutated precursor miRNA on the phenotype of each of said first and second cells;
wherein said phenotype is different, indicating that said loop sequence affects the role of said precursor miRNA; and
identifying said loop sequence as affecting the role of said precursor miRNA.
2. The method according to claim 1 , wherein said first and second cells are T-cells and said phenotype is the differentiation of said T-cells.