Self-cleaving ribozymes and uses thereof

Abstract

In certain embodiments, the disclosure relates to compositions and methods relating to a ribozyme-based gene regulation system that functions in mammalian cells. In certain specific embodiments, the disclosure relates to schistosome self-cleaving RNA mutant motifs.

1 - 13 . (canceled) 
     
     
         14 . A nucleic acid encoding a schistosome ribozyme mutant, wherein the schistosome ribozyme mutant comprises a loop on stem III and an aptamer in a position such that the cleaving activity of said schistosome ribozyme mutant can be modulated by binding of an effector to the aptamer. 
     
     
         15 . A host cell comprising the recombinant nucleic acid of  claim 14 . 
     
     
         16 . A recombinant polynucleotide molecule comprising:
 (a) a promoter;   (b) a nucleic acid encoding a nucleic acid product; and   (c) a nucleic acid encoding a schistosome ribozyme mutant motif,   wherein the nucleic acid of (b) and the nucleic acid of (c) are operably linked to the promoter and transcription of the nucleic acid of (b) and the nucleic acid of (c) produces a RNA molecule comprising said schistosome ribozyme mutant motif and a mRNA encoding said nucleic acid product, wherein said schistosome ribozyme mutant motif is capable of cleaving said RNA molecule intramolecularly.   
     
     
         17 . The polynucleotide molecule of  claim 16 , wherein the nucleic acid of (c) further comprises a nucleic acid encoding an aptamer which is at a position such that the cleaving activity of the schistosome ribozyme mutant motif is regulatable by binding of an effector to the aptamer. 
     
     
         18 . The polynucleotide molecule of  claim 16 , wherein the nucleic acid of (c) encodes at least two schistosome ribozyme mutant motifs. 
     
     
         19 . The polynucleotide molecule of  claim 16 , wherein the self-cleaving RNA mutant motif includes a loop on stem III. 
     
     
         20 . The polynucleotide molecule of  claim 19 , wherein the loop comprises at least three nucleotides. 
     
     
         21 . The polynucleotide molecule of  claim 19 , wherein the loop is selected from the group consisting of: 5′-UUCG-3′, 5′-CUUCGG-3′, and 5′-GCUUCGGU-3′. 
     
     
         22 . The polynucleotide molecule of  claim 19 , wherein the nucleotide C at position 7 is substituted with a nucleotide selected from the group consisting of: U, A and G. 
     
     
         23 . The polynucleotide molecule of  claim 19 , wherein the polynucleotide molecule is present in the genome of a cell. 
     
     
         24 . The polynucleotide molecule of  claim 19 , wherein the polynucleotide molecule is present on a vector. 
     
     
         25 . A host cell comprising the polynucleotide molecule of  claim 16 . 
     
     
         26 . The host cell of  claim 25 , wherein the cell is a mammalian cell. 
     
     
         27 . The host cell of  claim 25 , wherein the cell further comprises an inhibitor of the self-cleaving RNA mutant motif. 
     
     
         28 - 40 . (canceled) 
     
     
         41 . A method for modulating expression of a nucleic acid product in an individual, comprising:
 (a) obtaining cells from the individual under conditions appropriate for cell growth and cell division;   (b) introducing into said cells a DNA construct which comprises:
 (1) a promoter; 
 (2) a nucleic acid encoding a nucleic acid product; and 
 (3) a nucleic acid encoding said self-cleaving schistosome RNA mutant motif, wherein the nucleic acid of (2) and the nucleic acid of (3) are operably linked to the promoter, and transcription of the nucleic acid of (b) and the nucleic acid of (c) produces a RNA molecule comprising said self-cleaving schistosome RNA mutant motif and a mRNA encoding said nucleic acid product, wherein said self-cleaving schistosome RNA mutant motif is capable of cleaving said RNA molecule intramolecularly; 
   (c) returning said cell produced in step (b) to the individual; and   (d) administering to the individual an agent which modulatets cleavage of said self-cleaving schistosome RNA mutant motif such that expression of the nucleic acid product is modulated.   
     
     
         42 - 71 . (canceled)