RNA expression modulating or editing method via regulation of Cas13 protein
AssigneeKorea Advanced Institute of Science and Technology
InventorsWon Do Heo, Jeonghye Yu
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Abstract
The present invention relates to a RNA expression modulating or editing method through chemical or optogenetic regulation of Cas13 protein activity. Specifically, in order to regulate the activity of Cas13 in the CRISPR-Cas13 system, a fragment of the Cas13 protein was generated to enable recombination, and a chemogenetically or optogenetically bindable protein was linked to each fragment. And, it was confirmed that the Cas13 protein can be activated by treatment of a low molecular weight compound or irradiation of light. Accordingly, the method of the present invention can be effectively used for disease treatment by regulating RNA expression or editing RNA mutations related to disease.
Claims12
A method for regulating the activity of Cas13 protein, comprising a step of treating a first fragment containing the N-terminal segment of Cas13 protein and a first regulatory protein;
- and a second fragment containing a second regulatory protein and the C-terminal segment of Cas 13 protein with a compound;
- wherein the N-terminal segment of the Cas13 protein is a segment from the N-terminus of the Cas 13 protein to the 272nd, 286th, 351st, 480th or 928th amino acid of the Cas13 protein, and wherein the C-terminal segment of the Cas13 protein is a segment from the 273rd, 287th, 352nd, 481st or 929th amino acid of the Cas13 protein to the C-terminus of the Cas13 protein.
A method for modulating the expression of a target RNA comprising the following steps:
- 1) preparing a vector comprising a construct containing a sequence encoding the N-terminal segment of Cas13 protein and a sequence encoding a first regulatory protein;
- and a vector comprising a construct containing a second fragment comprising a sequence encoding a second regulatory protein and a sequence encoding the C-terminal segment of Cas13 protein;
- 2) injecting the vector above and crRNA (CRISPR RNA) capable of hybridizing to the target RNA sequence into mammalian cells;
- and
- 3) treating the cells with a compound;
- wherein the N-terminal segment of the Cas13 protein is a segment from the N-terminus of the Cas13 protein to the 272nd, 286th, 351st, 480th or 928th amino acid of the Cas13 protein, and
- wherein the C-terminal segment of the Cas13 protein is a segment from the 273rd, 287th, 352nd, 481st or 929th amino acid of the Cas13 protein to the C-terminus of the Cas13 protein.
Dependencies are inferred from the claim text.